# data[].attributes.headline
CRISPR-Cas9 Genome Editing: Weighing the Pros and Cons

# data[].attributes.subject
CRISPR-Cas9 Genome Editing: Weighing the Pros and Cons

# data[].attributes.body
Main idea:
Advantages and disadvantages of CRISPR.

Arguments/facts:
Advantages:
- CRISPR is useful in curing genetic diseases.
- It's cheap and also easy to control and to use.
- It can find the drugs in our body
Disadvantages:
- It's not always efficient and makes our cells be an unnormal cells.
- Needs time to produce it.

Dialogue:
- In which way, it can hurt our body?
- Why do people want to use CRISPR when needs time to create?
- It has cancer risk and other diseases risks.
- If the change and the modification on the DNA has not been done correctly are they responsible for us?
- The people who invented this should have a lot of responsibility and consequence for their clients because every moment could happen something to the clients so they should have a plan b helping them.

Summary:
CRISPR is an invention quite useful to change and to modify our genes, DNA, and other things. But not always is useful because it can be dangerous for our body and hurt us.
It's cheap and easy to control, can cure lots of diseases mostly cancer, and helps us with nutrition challenges. It also has a really good action that it can find the drugs in our body.
After editing your DNA your cells may not behave normally and correctly so that means that is not really efficient. Consumes a lot of time, needs a lot of responsibility.
So if you are thinking of having a change in your DNA you should think seriously and a lot of time so you know what you are doing with your body and that it could affect yourself and your next generation.

Smriti Tiwardi, (November 21, 2020)

11 Must-Know Advantages and Disadvantages of CRISPR Retrieved from https://honestproscons.com/advantages-and-disadvantages-of-crispr/

# data[].attributes.attachment_link.title
11 Must-Know Advantages and Disadvantages of CRISPR

# data[].attributes.attachment_link.description
Genome editing is an immensely effective technique that can assist you with your study to address a multitude of concerns. Here is a discussion of the key advantages and disadvantages of CRISPR and the process involved with genome editing using CRISPR.

# data[].attributes.headline
What Is CRISPR Currently Being Used For?

# data[].attributes.subject
What Is CRISPR Currently Being Used For?

# data[].attributes.body
Main idea:
CRISPR's potential and use
Arguments/facts:
-  some scientists are using CRISPR to potentially treat multiple myeloma, a cancer of the blood and bone marrow.
-  It is also being used to develop treatments for diseases like sickle cell anemia.
- it could potentially unintentionally edit the wrong gene.
- CRISPR is efficient, but it’s also almost irreversible, so if the strand doesn’t come back together perfectly, it can change the genetic code so potentially rendering it useless.

Dialogue:
-  "But even if CRISPR-Cas9 worked as planned, we simply don’t know the unintended consequences of altering this gene in humans" (Michael W. Richardson)

- could every baby in the future be edited to be immune to hereditary conditions and would it be ethical?
- without knowing the full consequences of any changes can we use CRISPR?

Summary:
Humans have been tinkering with genetics for thousands of years, even when we didn’t know it. But in the modern day, we can manipulate genes by means of CRISPR cas9. Which is an endlessly adaptable form of gene manipulation. The idea is basically that Cas9 could be reprogrammed to target, destroy, or replace specific genetic sequences.It is also being used to develop treatments for diseases like sickle cell anemia, possibly in the future CRISPR could be incredibly helpful for hereditary conditions such as the Huntington’s disease

What is CRISPR currently being used for?

With CRISPR you can directly observe what traits or behaviours are affected in lab mice, so CRISPR is used for studies but there are also clinical trials using CRISPR to treat several types of diseases and disorders.

Richardson, M. W. (2019, July 15)

What Is CRISPR Currently Being Used For?

Retrieved from https://www.brainfacts.org/In-the-Lab/Tools-and-Techniques/2019/CRISPR-Explained-071519

# data[].attributes.attachment_link.readable_text
Humans have been tinkering with genetics for thousands of years, even when we didn't know it. We created modern corgis and pomeranians by breeding pairs of wild wolves for docility, unknowingly selecting the genes controlling for size and temperament. Bananas became a breakfast staple as we continually bred its unappetizing ancestors to produce fruit with the least inedible seeds. It took centuries, but the results speak for themselves.

In the modern day, we manipulate individual genes in organisms, transforming a process taking generations into something that takes a fraction of the time. Scientists could breed a black mouse with a white mouse, locate and eliminate the genes for white fur in the mouse embryo, and rest assured that the baby mouse would grow black fur. Before the year 2000, researchers employed many different technologies to rewrite the genetics of organisms like this. But they were complex, expensive, and specific to a given plant or animal.

Enter CRISPR-Cas9. This gene-editing technique is based on the natural defense mechanism found in some bacteria. It uses a specific enzyme - Cas9 - to identify and eliminate predetermined genes and DNA sequences. It is a cheaper, more effective, and endlessly adaptable form of gene manipulation, and it seems to work in every model organism. Since its initial discovery around the turn of the millennium, scientists haven't fully realized the impact this technology may have on the field of genetics.

But how does it work, and what kind of treatments can we expect from CRISPR in the future? We asked Sean Ryder, professor of biochemistry and molecular pharmacology at UMass Medical School about this revolutionary breakthrough in genetics.

Where did the idea for CRISPR come from?

In the 1980s, Japanese scientist Yoshizumi Ishino and his team at Osaka University discovered clusters of DNA in some bacteria that contained strange repeats. It wasn't known why they repeated or what the clusters did, if anything. In 2001, researchers Francisco Mojica and Ruud Jansen coined a name for these repeating sections - CRISPRs, which stands for Clustered Regularly Interspaced Short Palindromic Repeats. A few years later, several independent groups of researchers noticed that these repeating clusters bracketed DNA that came from viruses that had attacked the bacteria before.

They realized that this bacteria was incorporating segments of DNA from viral invaders into its own genome and using it as an early warning system against attacks by the same virus. The CRISPR segments identify and flag the matching DNA in the viruses. But it's particular enzymes, including one called Cas9, that serve as weapons against the invaders. Cas9 carries a copy of the CRISPR sequence with it, searching for matching DNA in the viruses. Wherever it finds that matching DNA, Cas9 severs it and breaks it apart. Together, they form the CRISPR-Cas9 system.

CRISPR and Cas9 could have just been a clever defense mechanism for bacteria. But in 2012, researchers Jennifer Doudna, Emmanuelle Charpentier, and a separate team led by Lithuanian biochemist Virginijus Siksnys discovered that Cas9 could be reprogrammed to target, destroy, or replace specific genetic sequences, and not just in bacteria and viruses. That's when its potential for editing genes really appeared.

How does the technology work?

In bacteria, Cas9 carries crRNA - the genetic information of viruses to identify where to make their cuts. Researchers found that they could feed the enzyme fake genetic information, replacing the naturally occurring CRISPRs with an RNA sequence of their choosing, and send Cas9 to attack any gene they liked.

istock.com/traffic_analyzer

If Cas9 is assigned a specific RNA sequence and delivered to cells, it will hunt down corresponding sequences in the cellular DNA housed in the nuclei and perform a double-strand cut, severing the entire helix at a predetermined point. Severed DNA is an emergency for the cell, so it will immediately try to repair it. The simplest way it does this is by jamming the severed strands back into place and gluing them together. It's efficient, but it's also error-prone. And if the strand doesn't come back together perfectly, it can change the frame of the genetic code, potentially knocking out the gene that has been cut, or rendering it useless. Cas9 can also effectively cut out whole genes by using two guides that cut in two places, removing the intervening sequence, and the strands on either end will be glued together without the cut sequence.

It is possible to use a DNA sequence designed in the lab to repair a gene that has been cut by Cas9. This can be used to repair a broken gene, or to modify its function. However, this technology is less efficient than simply knocking out a gene.

What is CRISPR currently being used for?

In the laboratory, researchers regularly use CRISPR to alter genes in plant, bacteria, and animal models. If you knock out a particular gene in lab mice, you can directly observe what traits or behaviors are affected. This is how a lot of genetic research has been done for decades, but CRISPR makes these studies cheaper, faster, and more reliable.

There are also clinical trials using CRISPR to treat several types of diseases and disorders. For example, at the University of Pennsylvania, researchers are using CRISPR to potentially treat multiple myeloma, a cancer of the blood and bone marrow. They have harvested cells from bone marrow and edited the T cells, part of the immune system, to more accurately target cancer cells before putting them back into the body. The same technique has been used for sarcoma, a similar cancer. We don't have any results yet, but there is a lot of interest in this approach.

CRISPR-Cas9 is also being used to develop treatments for diseases like sickle cell anemia. In sickle cell anemia, having two bad copies of the Beta-globin causes severe symptoms, while having only one bad copy produces far fewer symptoms. So, researchers have harvested blood cells, run the CRISPR system to repair the one bad copy, and reintroduced the fixed cells back into the bloodstream. These techniques are clever, and they could have amazing ramifications for patients.

Which brain diseases and disorders could CRISPR treat in the future?

CRISPR will probably be most useful for hereditary conditions, such as Huntington's disease. In Huntington's, genes from one parent will always cause problems because it is a dominant gene, and there are no reliable treatments currently available. If you can knock out that dominant gene, the body will turn to the recessive, healthy gene for instructions instead. In this case, we can immediately identify an issue from the point of conception and try to act quickly to eliminate that dominant gene.

Hereditary conditions are the most likely targets for gene therapy because CRISPR, while more effective than previous technologies, still doesn't have a 100 percent success rate. The technology is always more effective in vitro because every cell in your body contains DNA, and a treatment may require the vast majority of those DNA strands to be altered. The fewer cells to edit, the more likely that CRISPR can effectively knock them out. If we know that there is likely to be a genetic problem early on, effective treatment is more likely.

There are still a lot of questions and hurdles that need to be overcome before we can edit entire human genomes. For example, a group of Chinese scientists used CRISPR to edit the genomes of twin girls to have an immunity to a certain strain of HIV. Editing was done during ICSI, intracytoplasmic sperm injection, and in vitro fertilization. Once it was determined that the embryos fertilized in this approach were modified, they were implanted into the mother and carried to term. That made them the first gene-edited children in history. Unfortunately, the work was performed in secret, with little oversight or medical rationale. CRISPR-Cas9 was used to destroy a normal gene in hopes of preventing the possibility of HIV infection. While no data has been published, analyses of information released during the Hong Kong genome editing conference revealed that both children contain at least two different modifications, and it's not clear whether all cells are modified.

But even if CRISPR-Cas9 worked as planned, we simply don't know the unintended consequences of altering this gene in humans. We don't know everything about the human genome. There's evidence the gene that was edited to prevent this particular strain of HIV also plays a role in intelligence and memory in animals. These girls may have had their cognitive abilities altered in some way. And there's evidence that the knocked-out gene helped protect against West Nile virus, so they may be more likely to contract that terrible disease in the future. Lastly, and perhaps most disturbing, the modifications could have unintended consequences, as a recent report has shown one particular mutation of CCR5 in humans is deleterious.

I find it extraordinarily risky and dangerous to create mutations in genes we don't fully understand, without knowing the full consequences of any changes. And when these girls grow up, they could pass on their edited genes. So, it's not just affecting the patient - it's affecting their progeny, and their progeny, down the entire family line. In order for me to be comfortable with this kind of gene editing, the technology will have to become more accurate, easier to apply, and capable of precisely editing genes, not just cutting or knocking them out. And I don't think we're there yet.

About the Author

# data[].attributes.attachment_link.description
Humans have been tinkering with genetics for thousands of years, even when we didn't know it. We created modern corgis and pomeranians by breeding pairs of wild wolves for docility, unknowingly selecting the genes controlling for size and temperament. Bananas became a breakfast staple as we continually bred its unappetizing ancestors to produce fruit with the least inedible seeds.

# data[].attributes.headline
Benefits and Ethical Concerns of CRISPR – Pros and Cons

# data[].attributes.subject
Benefits and Ethical Concerns of CRISPR – Pros and Cons

# data[].attributes.body
Main idea:

Benefits and ethical concert about CRISPR.

Arguments/facts:
- Pros:
- It can cure diseases by eliminating the bad cells for our body, they also found out new ways to kill the cancer cells.
- Cheap, useful.
- Cons:
- Changes the germ of our body, and maybe could pass to the next generation.
- Hurt our body.

Dialogue:
- What will happen if we removed all DNA from our body and CRISPR replaced them?
- If CRISPR is going to affect our body after curing it what will happen and how we will be treated.
- When we are modifying our DNA occurs something to our body will we die?

Summary:
There are benefits and not about the CRISPR.
Some benefits are cheap, easy to control, and more advanced than before, and that is another invention. It also is good for our body because it can eliminate the bad cells and the bad thing from our body so we can be healthier, it also treats diseases that are affecting lots of people like cancer.
The change of our body is affecting our next generation so it's dangerous.

Sara Guillan in Genetics

Benefits and Ethical Concerns of CRISPR – Pros and Cons. Retreview it from https://explorebiotech.com/crispr-pros-and-cons/

# data[].attributes.attachment_link.readable_text
CRISPR has become one of the most powerful gene-editing tools today. Unlike other genetic engineering tools, CRISPR is cheap, relatively easy to use and precise. Undoubtedly, its popularity has surged amongst scientists in the biotechnology industry.

Pros and Cons of CRISPR

While the benefits of CRISPR range from curing genetic conditions to organ transplants, ethicists fear its use in promoting desired traits rather than life-saving traits such as intelligence that could have long-term implications.

Benefits of CRISPR [Pros]

CRISPR technology has wide-ranging applications in various industries. Some of the benefits are discussed below.

Cancer Therapeutics: New immunotherapies can be developed using CRISPR to treat cancer. Scientists can genetically modify T-cells using CRISPR to locate and kill cancer cells.

Curing Genetic Diseases: CRISPR technology can eliminate the genes that cause genetic diseases such as diabetes, cystic fibrosis. Given that the scientific community has already mapped many genes that cause genetic disease, CRISPR could be used to cure faulty genes that cause genetic diseases.

Drug Research: Scientists are predicting that CRISPR could potentially speed up the drug discovery process, given the technology is relatively cheap, precise, and simple to use. Some of the drug makers in the world are already incorporating CRISPR technology into their drug research and discovery phase.

Pest Resilient Crops: According to Jennifer Doudna, CRISPR pioneer, genome editing could address pest and nutrition challenges facing agriculture, especially in light of climate change and rapid population growth. Earlier in 2018, USDA decided not to regulate about a dozen crops edited with CRISPR as GMOs partially due to the accuracy of the technology.

Ethical Concerns of CRISPR [Cons]

Changes to the Germ-line Cells: Genetically modifications to human embryos and reproductive cells such as eggs and sperms are called germline editing. Changes to the germline can be passed to the next generation. If the desired traits such as intelligence and strong muscles can be passed to the next generation, then how far are we from the designer babies?

Many countries have an outright ban on germline editing, whereas some countries do not have clear guidance. The United States and the United Kingdom are testing the water by considering exemptions in the case of monogenic hereditary disorders such as muscular dystrophy, cystic fibrosis, and Huntington's disease.

Bioweapons: The former U.S. director of national intelligence James Clapper listed "genome editing" among six threats in the section on weapons of mass destruction in 2016. Compared to other genetic engineering tools, CRISPR technology is relatively inexpensive and simple, which could make it attractive to terrorist organizations. The technology can be used to genetically modify bacteria or viruses to wage biological attacks against humans.

DARPA, US's secretive Defense Advanced Research Projects Agency, announced to invest US$65 million in 2017 over four years in seven teams that will investigate ways to make gene editing technologies safer and targeted. The program relates to both intentional and unintended consequences of gene editing technologies. Three main objectives of the program are

to develop processes that allow greater control of genome editing in living systems

to develop countermeasures that protect genome integrity in populations

and to investigate a way to remove engineered genes from living systems

Also Read - Pros and Cons of Having a Designer Baby

# data[].attributes.attachment_link.title
Benefits and Ethical Concerns of CRISPR - Pros and Cons

# data[].attributes.attachment_link.description
CRISPR has become one of the most powerful gene-editing tools today. Unlike other genetic engineering tools, CRISPR is cheap, relatively easy to use and precise. Undoubtedly, its popularity has surged amongst scientists in the biotechnology industry.

# data[].attributes.body
Main idea:
Is it safe and ethical to edit genes?
Arguments/facts:
-  Scientists are trying to take the genes from a long-extinct mammoth and add those to the Asian elephant to make it resistant to climate change.
- A researcher in China edited genes in embryos of twins to try to make them resistant to HIV and AIDS

Dialogue:
- Should we, as humans, be able to create and edit other humans to our liking? (for ex: designer baby)
- Should humans have this kind of power to create and edit species? neglecting natural evolution and stuff?

Summary:
Scientists have recently discovered that CRISPR has the potential to wipe out or rearrange large chunks of DNA, which could possibly turn off the wrong genes (the good genes) or trigger new diseases such as cancer. Also how far do people want to eradicate such “errors” in genes? Does that only for example include muscle sicknesses and cancer or does that also include dwarfism and blindness? People can also think of those things as unique and diverse.

Bess, M [Above the Noise]. (2019, April 4). Gene Editing & CRISPR: How Far Should We Go? Retrieved from  https://www.youtube.com/watch?v=wnlJ6dRfPFg

# data[].attributes.attachment_link.title
Gene Editing & CRISPR: How Far Should We Go?

# data[].attributes.body
Main idea:
CRISPR treatments safety
Arguments/facts:
- CRISPR cas9 could open an entire new world
- it could cure diseases

Dialogue:
- Would it be ethical to edit something or someone's genes so that they could never have that disease again?
- Can such a powerful tool cause harm to a healthy body?

Summary:
Using the technology of CRISPR cas9 a whole world of possibilities opens before you, but how safe and ethical is it actually?

There are many shady molecules out there that you’d never want to put in a person but are incredibly useful to help discover new biology. Does that mean we’re allowed to do that? The editing of DNA while having a disease can also for example damage the tumour suppressors and lead to cancer. And as with any disease, the safety of gene-editing therapy is all about the symptoms.

Corn, J. (2016, August 30). Safety for CRISPR. Retrieved from https://innovativegenomics.org/blog/safety-for-crispr/

# data[].attributes.attachment_link.readable_text
This post is all about establishing safety for CRISPR gene editing cures for human disease. Note that I did not say this post is about gene editing off-targets. We'll get there, but you might be surprised by what I have to say.

Contrary to what some might say or write, most of us gene editors do not have our heads in the sand when it comes to safety. From a pre-clinical, discovery research point of view, the safety of a given gene editing technology is relatively meaningless. There are many dirty small molecules out there that you'd never want to put in a person but are ridiculously useful to help unravel new biology. Given that pre-clinical researchers have the luxury of doing things like complementation/re- expression experiments and isolating multiple independent clones, let's dissociate everyone's personal research experience with CRISPR (all pre-clinical at this point) from questions of safety. Those experiences are useful and informative, but so far anecdotal and not necessarily tightly linked to the clinic.

Despite what we gene editors like to think, CRISPR safety is not a completely brand new world full of unexplored territory. While there are some important unanswered questions, there's a lot of precedent. Not only are other gene editing technologies already in the clinic, but non-specific DNA damaging agents are actually effective chemotherapies (e.g. cisplatin, temozolomide, etoposide). In the latter case, the messiness of the DNA damage is the whole point of the therapy.

Here are a few thoughts about the safety of a theoretical CRISPR gene editing therapy, in mostly random order. I'll preface this by saying that, while I have experience in the drug industry, I'm by no means an expert on clinical safety and defer to the real wizards.

Safety is about risk vs reward

Let's start with a big point: as with any disease, the safety of a gene editing therapy is all about the indication. The safety profile of a treatment for a glioblastoma (very few good treatment options for a fatal disease with fast progression) will be very different than a treatment for eczema. And the safety tolerance of a glioblastoma treatment that increases progression-free survival by only two days is going to look different than one that increases overall survival by five years. So there won't be One True Rule for gene editing safety, since most of the equation will be written by the disease rather than the therapy.

Gene editing safety is about functional genotoxicity

While most of the safety equation is about the disease, the treatment itself of course needs to be taken into account. At heart, gene editing reagents are DNA damaging agents, and so genotoxicity is a big concern. Does the intervention itself disrupt a tumor suppressor and lead to cancer? Does it break a key metabolic enzyme and lead to cell death? As mentioned above, there are plenty of DNA damaging agents that wreak havoc in the genome, but are tolerated because due to risk/reward and lack of a better alternative ( I'm especially looking at you, temozolomide). The key to this point is function of what gets disrupted.

With CRISPR, we have the marked advantage that guide RNAs tend to hit certain places within the genome. We know how to design the on-target and are still figuring out how to predict and measure the off-target. But even with perfect methods for off-targets, we'd still need to do the functional test. Consider a "traditional" therapeutic (small molecule or biologic) - while an in vitro off-target panel based on biochemistry is valuable, it's no substitute at all for normal-vs-tumor kill curves (as an example). And even those kill curves are no substitute for animal models. The long term, functional safety profile of a gene editing reagent is the key question, and with CRISPR I'd argue that we're still too early in the game to know what to expect. The good news is that ZFNs so far seem pretty good, giving me a lot of hope for gene editing as a class.

Gene editing safety is NOT about lists of sequences

You'd think that determining an exhaustive list of off-target sequences would be a critical part of any CRISPR safety profile. But in the example above, contrasting in vitro biochemical assays with organismal models, I consider lists of off-targets to be equivalent to the biochemical assay. I'm going to be deliberately controversial for a moment and posit that, for a therapeutic candidate, you shouldn't put much weight on its list of off-target sites. As stated above, you should instead care about what those off-targets are doing, and for that you might not even need to know where the off-targets are located.

When choosing candidate therapeutics in a pre-clinical mode, lists of off-target sequences can be very useful in order to prioritize reagents. If one guide RNA hits two off-target sites and another hits two hundred, you'd probably choose the former rather than the latter. But what if one the two off-targets is p53? What if the two-hundred are all intergenic? Given the fitness advantage to oncogenic mutations, the math involved in using sequencing (even capture-based technologies) to detect very rare off-target sites is daunting. Being able to detect a 1 in a million sequence-based event sounds incredible, but what if you need to edit as many as 20 million cells for a bone marrow transplant? That's twenty cells you might be turning cancerous but never even know it. Now we come right back around to function - you should care much more about the functional effect of your gene edit rather than a list of sequences. That list of sequences is nice for orders-of-magnitude and useful to choose candidate reagents, but it's no substitute for function.

Is gene editing safety about immunogenicity?

There are two big questions around gene editing immunogenicity: the immunogenicity of the reagent itself, and on-target immunogenicity if the edit introduces a sequence that's novel to the patient. What happens when the reagent itself induces a long-term immune response? For therapies that require repeat dosing, this can kill a program (hence a huge amount of work put into humanizing antibodies). A therapy that causes someone to get very sick on the second dose is not much good, nor is it useful if antibodies raised to the therapy end up blocking the treatment. But what about in situ gene editing?

Most of in situ gene editing reagents are synthetic or bacterial and so one might raise antibodies against them, but the therapy itself is (ideally), one-shot-to-cure. In that case, as long as there's not a strong naive immune response, maybe it doesn't matter if you develop antibodies to the editing reagent? There are few answers here for CRISPR, and most work with ZFNs has been with ex vivo edits, where the immune system isn't exposed to the editing reagent. Time will tell if this is a problem, and animal models will be key. Even more subtle, what happens when a gene edit causes re-expression of a "normal" protein that a patient has never before expressed (e.g. editing the sickle codon to turn mutant hemoglobin into wild type hemoglobin)?

The potential for a new immune response against a new "self" protein is probably related to the extent of the change - a single amino acid change (e.g. sickle cell) is probably less likely to cause problems than introducing a transgene (e.g. Sangamo's work inserting enzymes into the albumin locus for lysosomal storage disorders and hemophilia). But once again, I've heard a lot of questions and worry about this problem but very few answers. In vivo experiments are desperately needed, and the closer to a human immune system the better.

Moving forward based on the predictive data

As you've probably gathered by now, I have a healthy respect for functional characterization when it comes to safety. That's why it's absolutely critical that we keep moving forward and not let theoretical worries about arbitrary numbers of off-targets stifle innovation without data. These are tools that could some day help patients in desperate need and with few other options, so let the truly predictive functional data rule the day.

By Jacob Corn

# data[].attributes.attachment_link.description
This post is all about establishing safety for CRISPR gene editing cures for human disease. Note that I did not say this post is about gene editing off-targets. We'll get there, but you might be surprised by what I have to say.

# data[].attributes.body
Main idea:

The disease that CRISPR-Cas9 can cure and the future function of the CRISPR.

Arguments/facts:
- There are more than 8 diseases that CRISPR-Cas9.
- The diseases that it can cure are cancer, blood disorders, blindness, AIDS, cystic fibrosis, muscular dystrophy, Huntington's diseases, and covid-19.
- Each disease is cured with the help of the CRISPR, CRISPR is helping us to cure it so in the end, we need to totally cure ourselves of the disease.
- Cancer maybe not be cured totally with the CRISPR because it can passe things that we are not waiting for.

Dialogue:
- Why CRISPR-Cas9 can cure the covid-19? Even it can be left sequelae.
- If the modification makes the disease worst there will be more people that will change their DNA?
- Even though people say that CRISPR is precise, there's the science that can show that is not quite precise and safe.

Summary:
The CRISPR can cure 8 different diseases of our body.
Cancer is one of the diseases that can be cured by the CRISPR-Cas9.
In the first place was it started to cure cancer with the CRISPR-Cas9 was in China from 2019 to 2020 they were removing the gene that encodes for a protein called PD-1. After a university in the US add a substance into the treatment that China had. Blood disorders were adding the CRISPR into the bones, in the 5 patients, they didn't need anymore the blood of the patient to cure it. The blindness process happens inside the body. For AIDS there were many ways to cure it, the HIV was inserted into it so it can react. Cystic fibrosis is a disease about respiration so the CRISPR can help us to modify the original thing that caused the disease. Muscular dystrophy edits the proteins to be removed but in the end, is still functioning. In the end but not list Convid-19, this disease has been with us for the last 4 years and is cured with a new CRISPR called CRISPR/Cas13a that is invented by a scientist at Stanford University and its function is to stop creating inflammation in the lungs.
The technology of the CRISPR will be really advanced because what now it is doing and helping in the biggest diseases in the world is really proud and in the future, it will have better technology of the CRISPR.
Even though there are things that occur when you are modifying the DNA, could be not safe and not so precise as people say. So making a modification of the DNA is very serious because you can become and disabled person because of the unsafety in the modification.

Clara Rodíguez Fernandez, (September 13, 2021)

Eight Diseases CRISPR Technology Could Cure Retreview it from https://www.labiotech.eu/best-biotech/crispr-technology-cure-disease

# data[].attributes.attachment_link.readable_text
CRISPR technology offers the promise to cure any human genetic disease with gene editing; which one will be the first?
CRISPR-Cas9 was first used as a gene-editing tool in 2012. In just a few years, the technology has exploded in popularity thanks to its promise of making gene editing much faster, cheaper, and easier than ever before.

CRISPR is short for 'clustered regularly interspaced short palindromic repeats.' The term makes reference to a series of repetitive patterns found in the DNA of bacteria that form the basis of a primitive immune system, defending them from viral invaders by cutting their DNA.

Using this natural process as a basis, scientists developed a gene-editing technology called CRISPR-Cas9 that can cut a specific DNA sequence by simply providing it with an RNA template of the target sequence. This allows to then add, delete or replace elements within the target DNA sequence.

This system represented a big leap from previous gene-editing technologies, which required designing and making a custom DNA-cutting enzyme for each target sequence rather than simply providing an RNA guide, which is much simpler to synthesize.

CRISPR gene editing has already changed the way scientists do research, allowing a wide range of applications across multiple fields. But the technology could also hold great potential as a treatment for human diseases. In theory, CRISPR could let us edit any genetic mutation at will to cure any disease with a genetic origin. In practice, however, CRISPR is still in the beginning stages of its therapeutic development.

Here is a list of some of the first diseases that scientists are tackling using CRISPR-Cas technology, testing its possibilities and limits as a medical tool.

1. Cancer

China has been spearheading the first clinical trials using CRISPR-Cas9 as a cancer treatment. One of these studies was testing the use of CRISPR to modify immune T cells extracted from the patient. The gene-editing technology is used to remove the gene that encodes for a protein called PD-1. This protein found on the surface of immune cells is the target of some cancer drugs such as checkpoint inhibitors . This is because some tumor cells are able to bind to the PD-1 protein to block the immune response against cancer.

The trial tested this approach in 12 patients with non-small cell lung cancer at the West China Hospital. The results, published in April 2020, suggested the approach was feasible and safe.

However, a later article pointed out that the study revealed some of the technology's limitations, including variable efficiency in the genome-editing process. Some experts have recommended that the long-term safety of the approach remain under review. Others have suggested using more precise gene-editing approaches such as base editing .

In the US, a phase I trial run by the University of Pennsylvania tested the safety of a similar approach. The researchers used CRISPR to remove three genes that help cancer cells evade the immune system. They then added another gene to help the immune cells recognize tumors. T he revealed that the treatment was safe in patients with advanced forms of cancer.

Meanwhile, the company CRISPR Therapeutics is currently running a global phase I trial that is expected to recruit over 130 patients with blood cancer to test a CAR-T cell therapy made using CRISPR technology.

2. Blood disorders

The blood disorders beta-thalassemia and sickle cell disease, which affect oxygen transport in the blood, are the target of a CRISPR treatment being developed by CRISPR Therapeutics and its partner Vertex Pharmaceuticals.

The therapy consists of harvesting bone marrow from the patients and using CRISPR technology to make them produce fetal hemoglobin. This is a natural form of the oxygen-carrying protein that binds oxygen much better than the conventional adult form. The modified cells are then reinfused into the patient.

In December, preliminary results revealed that all five patients with thalassemia haven't required any blood transfusions since receiving the treatment , and the two patients with sickle cell disease have so far not experienced any of the usual bleeding episodes caused by their condition.

Hemophilia is another blood disorder that CRISPR technology could tackle, although development is still at the preclinical stage. CRISPR Therapeutics is working with Casebia on an where the gene-editing tool is delivered directly to the liver. Last year, Intellia Therapeutics and Regeneron Pharmaceuticals teamed up to pursue the development of hemophilia treatments based on genome editing.

3. Blindness

Many hereditary forms of blindness are caused by a specific genetic mutation, making it easy to use CRISPR-Cas9 to treat it by targeting and modifying a single gene. In addition, the activity of the immune system is limited in the eye, which can circumvent any problems related to the body rejecting the treatment.

The company Editas Medicine is working on a CRISPR therapy for Leber congenital amaurosis, the most common cause of inherited childhood blindness, for which there is currently no treatment. The treatment aims to use CRISPR to restore the function of light-sensitive cells before the children lose sight completely by fixing the most common genetic mutation behind the disease.

Last year the company started a phase I/II trial, with results expected by 2024. This is the first trial to test an CRISPR treatment, in which the gene editing happens directly inside the patient's body rather than on cells extracted from their body and then returned to it.

4. AIDS

There are several ways CRISPR could help us in the fight against AIDS. One is using CRISPR to cut the viral DNA that the HIV virus inserts within the DNA of immune cells. This approach could be used to attack the virus in its hidden, inactive form, which is what makes it impossible for most therapies to completely get rid of the virus.

Another approach could make us resistant to infections. Certain individuals are born with a natural resistance to HIV thanks to a mutation in a gene known as CCR5, which encodes for a protein on the surface of immune cells that HIV uses as an entry point to infect the cells . The mutation changes the structure of the protein so that the virus is no longer able to bind to it.

This approach was used in a very controversial case in China two years ago in which human embryos were genetically edited to make them resistant to HIV infections. The experiment caused outrage among the scientific community, with some studies pointing out that the 'CRISPR babies' might be at a higher risk of dying younger. The general consensus seems to be that more research is needed before this approach can be used in humans, especially as recent studies have pointed out this practice can have a of unintended genetic edits in embryos.

5. Cystic fibrosis

Cystic fibrosis is a genetic disease that causes severe respiratory problems. Although there are treatments available to deal with the symptoms, the life expectancy for a person with this disease is only around 40 years. CRISPR technology could help us get to the origin of the problem by editing the mutations that cause cystic fibrosis, which are located in a gene called .

Last year, researchers in the Netherlands used base editing to repair mutations in the cells of people with cystic fibrosis without creating damage elsewhere in their genetic code. In addition, companies such as Editas Medicine, CRISPR Therapeutics, and Beam Therapeutics have plans to develop treatments for cystic fibrosis using CRISPR systems.

Cystic fibrosis can be caused by multiple different mutations in the target gene, however, meaning that different therapies will have to be developed for different genetic defects. Editas Medicine has stated that it will be looking at the most common mutations, as well as some of the rare ones for which there is no treatment.

6. Muscular dystrophy

Duchenne's muscular dystrophy is caused by mutations in the gene, which encodes for a protein necessary for the contraction of muscles. Children born with this disease suffer progressive muscle degeneration, and existing treatments are limited to a fraction of patients with the condition.

Research in mice has shown CRISPR technology could be used to fix the multiple genetic mutations behind the disease. In 2018, a group of researchers in the US used CRISPR to cut at 12 strategic 'mutation hotspots' covering the majority of the estimated 3,000 different mutations that cause this muscular disease. A company called Exonics Therapeutics was spun out to further develop this approach.

Editas Medicine is also working on a CRISPR therapy for Duchenne's muscular dystrophy. The company is following a broader approach where instead of fixing specific mutations, CRISPR gene editing is used to remove whole sections of the mutated protein, which makes the protein shorter but still functional.

7. Huntington's disease

Huntington's disease is a neurodegenerative condition with a strong genetic component. The disease is caused by an abnormal repetition of a certain DNA sequence within the huntingtin gene. The higher the number of copies, the earlier the disease will manifest itself.

Treating Huntington's could be tricky, as any off-target effects of CRISPR in the brain could have very dangerous consequences. To reduce the risk, scientists are looking at ways to tweak the genome-editing tool to make it safer.

In 2018, researchers at the Children's Hospital of Philadelphia revealed a version of CRISPR-Cas9 that includes a self-destruct button . A group of Polish researchers opted instead for pairing CRISPR-Cas9 with an enzyme called nickase to make the gene editing more precise .

8. Covid-19

In the face of the pandemic, CRISPR has quickly been put to the use of making fast screening tests. In the longer term, the gene-editing tool might allow us to fight Covid-19 and other viral infections.

Scientists at Stanford University have developed a method to program a version of the gene-editing technology known as CRISPR/Cas13a to cut and destroy the genetic material of the virus behind Covid-19 to stop it from infecting lung cells . This approach has shown to reduce the viral load in human cells by 90% and to work against 90% of all existing and emerging coronaviruses.

Another research group at the Georgia Institute of Technology has used a similar approach to destroy the virus before it enters the cell. The method was tested in live animals , improving the symptoms of hamsters infected with Covid-19. The treatment also worked on mice infected with influenza, and the researchers believe it could be effective against 99% of all existing influenza strains.

The future of CRISPR technology

Considering that CRISPR-Cas9 is a relatively new development in the world of biology, research has only begun to scratch the surface of the role it could play in the future of medicine. The examples listed here are just the first attempts at using CRISPR technology as a therapy. As they progress, we can expect more and more indications to be added to the list.

One of the biggest challenges to turn this research into real cures is the many unknowns regarding the potential risks of CRISPR therapy. Some scientists are concerned about possible off-target effects as well as immune reactions to the gene-editing tool. But as research progresses, scientists are proposing and testing a wide range of approaches to tweak and improve CRISPR in order to increase its efficacy and safety.

Hopes are high that CRISPR-Cas9 technology will soon provide a way to target and destroy complex diseases such as cancer and AIDS, and even target genes associated with mental illnesses.
Images via NIH /Flickr; Shutterstock. This article was originally published in June 2018 and has since been updated to reflect the latest developments in CRISPR research.

# data[].attributes.attachment_link.title
Eight Diseases CRISPR Technology Could Cure

# data[].attributes.attachment_link.description
CRISPR technology offers the promise to cure any human genetic disease with gene editing; which one will be the first? CRISPR-Cas9 was first used as a gene-editing tool in 2012. In just a few years, the technology has exploded in popularity thanks to its promise of making gene editing much faster, cheaper, and easier than ever before.

# data[].attributes.body
Main idea:
CRISPR-Cas9 is a genome-editing tool by removing, adding or altering sections of the DNA sequence that is creating a buzz in the science world. It is faster, cheaper and more accurate than previous techniques of editing DNA and has a wide range of potential applications.

Arguments/facts:
- The CRISPR-Cas9 system consists of two key molecules that introduce a change (mutation) into the DNA. These are:

An enzyme called Cas9. This acts as a pair of molecular scissors that can cut the two strands of DNA at a specific location in the genome so that bits of DNA can then be added or removed.

A piece of RNA called guide RNA (gRNA). This consists of a small piece of pre-designed RNA sequence. It guides Cas9 to the right part of the genome and makes sure that the Cas9 enzyme cuts at the right point in the genome.

- The Cas9 follows the guide RNA to the same location in the DNA sequence and makes a cut across both strands of the DNA. The cell recognises that the DNA is damaged and tries to repair it.
- CRISPR-Cas9 has a lot of potential as a tool for treating a range of medical conditions that have a genetic component
- Carrying out gene editing in germline (reproductive) cells is currently illegal al in the UK and most other countries.

- Many of the proposed applications involve editing the genomes of somatic non-reproductive) cells but there have been a lot of interest in and debate about the potential to edit germline (reproductive) cells.
- Scientists are keen to find a way to ensure that the CRISPR-Cas9 binds and cuts accurately.

Dialogue:
- What if we cut the wrong piece of DNA?
- Not knowing the second effects of CRISPR, should we test it on humans?
- I think that it is still dangerous because we don't know what could happen 100%
- How do we inject the right amount of protein Cas-9 into our bodies?
- Could we really trust RNA?

Summary:
Scientists can use DNA repair machinery to introduce changes to one or more genes in the genome of a cell of interest using CRISPR-Cas9. They have discovered this by observing bacteria that have Cas9 protein, and have adapted this system to test it on other organisms. The CRISPR-Cas9 system currently stands out as the fastest, cheapest and most reliable system for editing genes. However, because any changes made in germline cells will be passed on from generation to generation it has important ethical implications. It is likely to be many years before CRISPR-Cas9 is used routinely in humans. Much research is still focusing on its use in animal models or isolated human cells. There is still a lot of work focusing on eliminating other targets CRISPR-Cas9 might cut that were not intended to be edited.

(2021, October 25). What is CRISPR-Cas9? Retrieved from https://www.yourgenome.org/facts/what-is-crispr-cas9

# data[].attributes.attachment_link.readable_text
CRISPR-Cas9 is a genome editing tool that is creating a buzz in the science world. It is faster, cheaper and more accurate than previous techniques of editing DNA and has a wide range of potential applications.

What is CRISPR-Cas9?

CRISPR-Cas9 is a unique technology that enables geneticists and medical researchers to edit parts of the genome by removing, adding or altering sections of the DNA sequence.

It is currently the simplest, most versatile and precise method of genetic manipulation and is therefore causing a buzz in the science world.

How does it work?

The CRISPR-Cas9 system consists of two key molecules that introduce a change (mutation) into the DNA. These are:

an enzyme called Cas9. This acts as a pair of 'molecular scissors' that can cut the two strands of DNA at a specific location in the genome so that bits of DNA can then be added or removed.

a piece of RNA called guide RNA (gRNA). This consists of a small piece of pre-designed RNA sequence (about 20 bases long) located within a longer RNA scaffold. The scaffold part binds to DNA and the pre-designed sequence 'guides' Cas9 to the right part of the genome. This makes sure that the Cas9 enzyme cuts at the right point in the genome.

The guide RNA is designed to find and bind to a specific sequence in the DNA. The guide RNA has RNA bases that are complementary to those of the target DNA sequence in the genome. This means that, at least in theory, the guide RNA will only bind to the target sequence and no other regions of the genome.

The Cas9 follows the guide RNA to the same location in the DNA sequence and makes a cut across both strands of the DNA.

At this stage the cell recognises that the DNA is damaged and tries to repair it.

Scientists can use the DNA repair machinery to introduce changes to one or more genes in the genome of a cell of interest.

Some bacteria have a similar, built-in, gene editing system to the CRISPR-Cas9 system that they use to respond to invading pathogens like viruses, much like an immune system.

Using CRISPR the bacteria snip out parts of the virus DNA and keep a bit of it behind to help them recognise and defend against the virus next time it attacks.

Scientists adapted this system so that it could be used in other cells from animals, including mice and humans.

What other techniques are there for altering genes?

Over the years scientists have learned about genetics and gene function by studying the effects of changes in DNA.

If you can create a change in a gene, either in a cell line or a whole organism, it is possible to then study the effect of that change to understand what the function of that gene is.

For a long time geneticists used chemicals or radiation to cause mutations. However, they had no way of controlling where in the genome the mutation would occur.

For several years scientists have been using 'gene targeting' to introduce changes in specific places in the genome, by removing or adding either whole genes or single bases.

Traditional gene targeting has been very valuable for studying genes and genetics, however it takes a long time to create a mutation and is fairly expensive.

Several 'gene editing' technologies have recently been developed to improve gene targeting methods, including CRISPR-Cas systems, transcription activator-like effector nucleases (TALENs) and zinc-finger nucleases (ZFNs).

The CRISPR-Cas9 system currently stands out as the fastest, cheapest and most reliable system for 'editing' genes.

What are the applications and implications?

CRISPR-Cas9 has a lot of potential as a tool for treating a range of medical conditions that have a genetic component, including cancer, hepatitis B or even high cholesterol.

Many of the proposed applications involve editing the genomes of somatic (non-reproductive) cells but there has been a lot of interest in and debate about the potential to edit germline (reproductive) cells.

Because any changes made in germline cells will be passed on from generation to generation it has important ethical implications.

Carrying out gene editing in germline cells is currently illegal in the UK and most other countries.

By contrast, the use of CRISPR-Cas9 and other gene editing technologies in somatic cells is uncontroversial. Indeed they have already been used to treat human disease on a small number of exceptional and/or life-threatening cases.

What's the future of CRISPR-Cas9?

It is likely to be many years before CRISPR-Cas9 is used routinely in humans.

Much research is still focusing on its use in animal models or isolated human cells, with the aim to eventually use the technology to routinely treat diseases in humans.

There is a lot of work focusing on eliminating 'off-target' effects, where the CRISPR-Cas9 system cuts at a different gene to the one that was intended to be edited.

Better targeting of CRISPR-Cas9

In most cases the guide RNA consists of a specific sequence of 20 bases. These are complementary to the target sequence in the gene to be edited. However, not all 20 bases need to match for the guide RNA to be able to bind.

The problem with this is that a sequence with, for example, 19 of the 20 complementary bases may exist somewhere completely different in the genome. This means there is potential for the guide RNA to bind there instead of or as well as at the target sequence.

The Cas9 enzyme will then cut at the wrong site and end up introducing a mutation in the wrong location. While this mutation may not matter at all to the individual, it could affect a crucial gene or another important part of the genome.

Scientists are keen to find a way to ensure that the CRISPR-Cas9 binds and cuts accurately. Two ways this may be achieved are through:

the design of better, more specific guide RNAs using our knowledge of the DNA sequence of the genome and the 'off-target' behaviour of different versions of the Cas9-gRNA complex.

the use of a Cas9 enzyme that will only cut a single strand of the target DNA rather than the double strand. This means that two Cas9 enzymes and two guide RNAs have to be in the same place for the cut to be made. This reduces the probability of the cut being made in the wrong place.

# data[].attributes.attachment_link.description
CRISPR-Cas9 is a genome editing tool that is creating a buzz in the science world. It is faster, cheaper and more accurate than previous techniques of editing DNA and has a wide range of potential applications. CRISPR-Cas9 is a unique technology that enables geneticists and medical researchers to edit parts of the genome by removing, adding or altering sections of the DNA sequence.

# data[].attributes.headline
10 Amazing Things Scientists Just Did with CRISPR

# data[].attributes.subject
10 Amazing Things Scientists Just Did with CRISPR

# data[].attributes.body
Main idea:
It's like someone has pressed fast-forward on the gene-editing field: A simple tool that scientists can wield to snip and edit DNA is speeding the pace of advancements that could lead to treating and preventing diseases. The tool used for this is called CRISPR-Cas9. Recent advances in the fights against 10 diseases that demonstrate CRISPR's capabilities and hint at things to come:

Arguments/facts:
- Cancer: Like many diseases, cancer results from a mutation in a person's genome, researchers say it's possible that a CRISPR-based treatment could one day slow the speed at which a tumour spreads, or perhaps reverse the disease completely.
- HIV: The virus infects the immune cells in the body that attack viruses, and it's also a notorious mutator. In May 2017, some researchers in a university in Pittsburgh used CRISPR to snip the virus from the cell it was infecting, shutting down the virus's ability to replicate.
- Huntington's disease: A fatal genetic disorder that causes nerves in the brain to deteriorate over time. In June 2017, scientists reported that they had reversed the disease in lab mice using CRISPR to snip out part of the mutant huntingtin gene that produces the toxic bits, which might help the condition.
- Duchenne muscular dystrophy: A debilitating condition that develops because of a mutation in a single gene, called the dystrophin gene, which is one of the longest genes in the body. Using a variation of the CRISPR tool, called CRISPR-Cpf1, corrected the mutation that causes Duchenne muscular dystrophy.
- It could prevent some birth blindness by fixing the genetic error that causes it.
- It could reduce chronic pain by altering genes to reduce inflammation using CRISPR.
- Malaria: Scientists have been trying to reduce malaria by also reducing mosquitoes. In December 2015, scientists had identified three genes to reduce female mosquito fertility. They also announced that they had found that CRISPR could work to target at least one of them.
- Lime disease: It is caused by a tick-borne bacterium that can spread from deer-tick bites to people. Scientists want to reduce the disease by using CRISPR-Cas9 to genetically modify white-footed mice in a way that would make them and their offspring become immune to the bacteria and unable to pass it along to ticks.
- CRISPR can also be used to modify the genomes of plants. Scientists are investigating ways to harness the tool's gene-editing ability to reduce disease in some crops and make others more robust.
- Editing human embryos: The scientists used CRISPR-Cas9 to introduce and then edit out disease-causing mutations from human embryos. The study was done to show that genetic editing could be done at the embryonic stage. The embryos were not implanted in a human.

Dialogue:
- Would we use it not only for therapeutic needs but for human physical traits too?
- Could CRISPR make twins?
- With all of this gene editing to cure diseases, maybe other stronger diseases will appear from mutations.
- If we knew about dangerous consequences, would we still do it to cure other diseases?
- Should we use Crispr on the human germline, so that modifications are inherited by future generations?

Summary:
Diseases such as cancer, HIV, blindness, chronic pain, muscular dystrophy, Huntington's disease, etc. have been a total challenge for everyone up until now. However, now, with CRISPR, scientists may have found a solution to cure these, including some other genetic related diseases. It can be done by cutting or replacing the genetic malfunctions that cause these.

Tracy, S. (2017, June 26). 10 Amazing Things Scientists Just Did with CRISPR. Retrieved from https://www.livescience.com/59602-crispr-advances-gene-editing-field.html

# data[].attributes.attachment_link.title
10 Amazing Things Scientists Just Did with CRISPR

# data[].attributes.attachment_link.description
A new tool called CRISPR is letting scientists cut and snip DNA in better ways, and has led to a slew of new research that touches on many human diseases.

# data[].attributes.headline
Genetic engineering will change everything forever - CRISPR

# data[].attributes.subject
Genetic engineering will change everything forever - CRISPR

# data[].attributes.body
Main idea:
Gene editing was extremely expensive, complicated and took a long time to do. This has now changed with a revolutionary new technology called CRISPR. The costs of engineering have shrunk by 99%. Instead of a year, it takes a few weeks to conduct experiments, and basically anyone with a lab can do it. CRISPR works for every type of cell and has the potential to change humanity forever.

Arguments/facts:
- Genetic engineering was first tested on plants and the earliest genetically modified animal (a mouse) was born in 1974. The first food modified in the lab went on sale in 1994: the Flavr Savr tomato. It's a tomato given a much longer shelf life.
- In 2015, scientists used CRISPR to cut the HIV virus out of living cells from patients in the lab, proving that it was possible.
- CRISPR can edit our immune cells and make them better cancer hunters in our bodies. This could be achieved by just getting a few injections. In 2016, the US approved cancer treatment with CRISPR and China approved lung cancer treatment with immune cells modified with CRISPR.
- Experimenting with human embryos has already happened in China in 2015 and 2016 and were partially successful on their second attempt.
- Modified humans could alter genomes of our entire species, passing their engineered traits to their children and generations ahead.

Dialogue:
- Could a state force gene editing on their subjects?
- What could stop an army of genetic engineered supersoldiers?
- How far would humanity go with CRISPR?
- Could we do CRISPR at home?
- How was CRISPR discovered?

Summary:
CRISPR offers the ability to edit live cells, switch genes on and off, and target and study particular DNA sequences.
Engineered humans might be better equipped to cope with high-energy food, eliminating many diseases like obesity. With possession of a modified human system, we might become immune to most diseases that are deathly nowadays.  ​We could extend our life expectancy by centuries, be used for reproductive purposes, and stop, slow down or even reverse ageing. Even further from the future, we could engineer humans to be equipped for extended space travel and cope with different conditions on other planets.

Kurzgesagt - In a Nutshell. (2016, August 10). Genetic engineering will change everything forever - CRISPR. Retrieved from https://youtu.be/jAhjPd4uNFY

# data[].attributes.attachment_link.title
Genetic Engineering Will Change Everything Forever – CRISPR

# data[].attributes.body
Main idea:

Experts debating the accomplish with the powerful DNA editing tool and what can it cause.

Arguments/facts:
- We can modify the DNA from an animal, plant, and human.
- It's cheap, simple to manage and it contains a substance that can kill bacteria.
- You can manage it by yourself at your house.
- You can be like you want after changing your genes.

Dialogue:
- Does it bring bad things to our body or does it hearts it?
- Why did they create the CRISPR if before there was another invention with the same function even was difficult and expensive? They could make it cheaper and easier to control.
- I'm worried about the people who want to change their body to a technological body.
- Could they have their non-electrical body after changing it to a technical body?

Summary:
CRISPR is a technic that has been used since a long time ago and is very useful.
The technic CRISPR is letting us add DNAs to any part of our body and this technic is also can be added to animals and plants. It's cheap to buy and is pretty easy to control or use, I think that is a good invention because it can kill the bacteria inside our body.
Before CRISP there was another invention called TALENS that was not cheap, not easy, not easy to control invention for the scientist.

Annie, S. (2017, November 2). Mail-Order CRISPR Kits Allow Absolutely Anyone to Hack DNA. Retrieved from https://www.scientificamerican.com/article/mail-order-crispr-kits-allow-absolutely-anyone-to-hack-dna

# data[].attributes.attachment_link.readable_text
"We aren't going to get sick, are we?" my roommate Brett asked me. He cringed as I knelt down and stuffed a plate of E. coli bacteria-which came as part of the DIY CRISPR-Cas9 kit I bought online-into our fridge next to cartons of eggs, strawberry jam, bottles of beer and a block of cheese.

"No, we won't. The label says 'non-pathogenic,'" I replied, trying to sound assuring. But honestly, I had no clue what I was doing. I nudged all the food up against the fridge wall, and left a two-inch border around the plate of living cells-a no man's land between the microbes and our dinner. A couple inches probably would not stop the bugs, but I figured it couldn't hurt.

CRISPR-Cas9 (or CRISPR, for short) has given scientists a powerful way to make precise changes to DNA-in microbes, plants, mice, dogs and even in human cells. The technique may help researchers engineer drought-resistance crops, develop better drugs, cure genetic disorders, eradicate infectious diseases and much more. Ask any biologist, and they'll likely tell you that CRISPR is revolutionary. It's cheap and effective, and in many cases, it works much better than older methods for making genetic modifications. Biologists will also tell you that CRISPR is very easy to use. But what does "easy to use" mean?

I am not a DIY scientist, much less a professional scientist. You won't find me swabbing my cheek cells for DNA or tinkering with yeast in a lab on the weekend. But I wondered: Is CRISPR so easy thateven amateurs like me can make meaningful contributions to science? And also, does this new technique make gene editing so accessible that we need to worry about DIY scientists cooking up pandemic viruses in their basements? If you Google 'DIY CRISPR,' stories such as "What Happens If Someone Uses this DIY Gene Hacking Kit to Make Mutant Bacteria?" pop up.

I attempted to find answers to all these questions myself, starting with the plate of bacteria in the kitchen of my San Francisco apartment.

CUT AND SPLICE

CRISPR stands for "clustered regularly interspaced short palindromic repeats." The CRISPR system is made up of two components: a protein called Cas9 and a guide RNA,a string of nucleic acid molecules with a certain genetic code. Put them together, and they create a tool you can use to tweak an organism's genome. To do this, CRISPR searches the organism's DNA for a certain sequence-specifically, the one encoded by the guide RNA, which holds the inverse sequence of your target DNA. "Cas9 opens up the DNA, it separates the strands of the double helix in a very small area, and allows the guide RNA to pair with one of the strands," explains Dana Carroll, a professor of biochemistry at the University of Utah. "If it is a good match, cutting occurs. If it is not a good match, Cas9 and [the] guide RNA fall off and try again somewhere else." When it finds the right sequence, the Cas9 protein slices the DNA at that precise spot.

At this point, if you leave the cell alone, it will usually mend CRISPR's cut-but it will occasionally also make a mistake in the repair process, breaking a gene or other parts of the genome. Since CRISPR repeatedly goes back and slices the DNA again after the cell mends it, the gene eventually breaks, or, in technical terms, gets knocked out. And, if you add new DNA, the cell may incorporate it while fixing the cut. This means you can insert DNA where you want to in the genome-you just need to know the organism's genetic sequence of your desired target area.

Scientists originally discovered this sophisticated system in archaea and bacteria, which deploy CRISPR to chop up invading viruses. But a few years ago, researchers figured out how to repurpose CRISPR to run inpretty much any living thingthey want. And now it's made genetic engineering easier than ever before.

For my own experiment, everything I needed came in a small cardboard box-an assortment of bottles, tubes, plates, powders and liquids (plus E. coli). I ordered my kit for $130 from the crowd-funding site Indiegogo as part of a campaign created by Bay Area biohacker Josiah Zayner. Zayner has a PhD in molecular biophysics and spent two years as a research fellow at NASA. He ran the crowdfunding campaign out of his apartment, and by the end of it, he had raised over $70,000 and sold 250 DIY CRISPR kits-one of which now sat on my kitchen table. Zayner has now soldover a thousand kits, largely on his company's website, The Odin.

The idea behind the kit experiment is quite simple. The goal: modify the E. coli so that it can grow on an antibiotic called streptomycin, which normally kills bacteria. With materials and instructions from the kit, I will introduce CRISPR into the bacteria cells, and use it to rewrite a tiny part of their DNA, creating genetically altered cells that happily thrive on streptomycin. In the end, CRISPR will track down and then change only a single base pair (which are the building blocks for DNA)out of the 4.6 million base pairs in the E. coli genome. It will swap out the chemical compound adenine for cytosine-or, in terms of the genetic alphabet, an "A" for a "C." Because of that tiny code change, my bacteria cells will make the amino acid lysine instead of another one, threonine. If my gene editing succeeds, this will stop streptomycin from interfering with the E. coli.

On a Monday afternoon in May, I donned latex gloves and spread newspaper over the dinner table. I grabbed three small plastic tubes from the freezer, picked up a pipette-a hand-held instrument, used in labs to measure out liquids-from my DIY kit, and started adding the CRISPR ingredients to tubes of E. coli.

I put in the Cas9 protein and guide RNA,which came as liquids in small, plastic test tubes,to my bacteria cells. Then I dipped my pipette into a small tube of DNA and tried to suck up 10 microliters. Nothing came out. I squinted at the tiny drop of clear liquid, and realized it had frozen solid. I had no clue why. Uh oh, I hope that's not a problem. Would it harm my experiment? I had absolutely no idea, so I just waited for the DNA solution to thaw and then squirted it into the bacteria tube. After several more steps, I spread my CRISPR'd bacteria onto three plastic plates and put them in my laundry room. The instructions said to wait 24-48 hours, then check for small white dots of bacteria. If I saw dots, CRISPR had done its job of splicing in the streptomycin-resistance gene. If not ... well, failure is also part of the scientific process.

I had no problem conducting the experiment- CRISPR is easy, I concluded. I basically just measured, scraped and stirred a bunch of ingredients, occasionally cooling them or heating them up. But for all the godlike powers that I imagined CRISPR gave me, I actually had little say over what I did to my bacteria. Everything was predetermined, with instructions laid out for me like steps in a cookbook: "Add 100 microliters Transformation mix to a new centrifuge tube," "Incubate this tube in the fridge for 30 minutes," and so on. Ultimately, I had made zero decisions. Of course, I could have designed a custom-made CRISPR experiment-but it would have taken more time, more materials, more money, and a lot more knowledge than I currently had.

I inspected my bacteria 48 hours later. Crossing my fingers, I lifted the lid on the first plate. No white dots. Then the second plate: nothing. My stomach sank with disappointment. Then I gently raised the lid off the third plate, and saw ... something. The plate had two faint milky white circles. Had CRISPR worked? Maybe. But then why did one plate have white spots, but not the other plates? I'd followed the same steps for each of them. Maybe my mind was playing tricks on me. Or perhaps I had contaminated the third plate. If only I could show my plates to someone who knew how to interpret them. Like a scientist. Unfortunately, there was no one to ask in my kitchen.

OPEN TO EVERYONE

A few weeks later, I drove 40 miles south of San Francisco to meet a DIY scientist named Johan Sosa, who knows way more about CRISPR than I do. We met at BioCurious, a community laboratory in Sunnyvale where he works most weekends and some evenings. Located in Santa Clara, BioCurious is a co-working space outfitted with science equipment and shared by "scientists, technologists, entrepreneurs, and amateurs who believe that innovations in biology should be accessible, affordable, and open to everyone," according to its Web site.The lab is funded by donations and members-Sosa is one of itsseveral dozenmembers.

At six foot, five inches, Sosa towers over most people. "I'm probably the tallest DIY biologist," he jokes. He laughs easily, which offsets his height to lend him a gentle, laid-back manner. He is 40 years old, his dark hair flecked with silver. Originally from Sri Lanka, Sosa came to the U.S. for college at 15 years old to study computer science, and he's worked for Bank of America and IBM as a computer security specialist and software engineer. He now has a day job in computer security-but he spends most of his free time at BioCurious. "You could say I have no life," he chuckles, "This is my biggest hobby." He has learned everything he knows about science (both theory and lab techniques) from others at BioCurious, by reading science papers, watching YouTube videos, attending lectures, and also through trial-and-error in his own research.

Sosa is one of a few DIY scientists at BioCurious using CRISPR. He first read about the technique in 2012, from one of biochemist Jennifer Doudna'spapers in the journal Science. Doudna, a professor at University of California, Berkeley, is one of the pioneers of CRISPR. "I didn't think it was that big of a deal, because I knew there were already other ways of modifying DNA," he recalls, "But I did think, 'This is something I could do.'" He started experimenting with CRISPR in 2013.

On a muggy, overcast afternoon, I followed Sosa inside BioCurious. We strolled through a lobby and into a large windowless room. It had big cabinets crammed with bottles of liquid, racks of latex gloves, a giant bio-hood, microwave and fridge. Microscopes, scales, centrifuges and a bunch of other well-worn science equipment lay scattered across lab desks. A calm thrum of buzzing machinery filled the room, and test tubes shook quietly in an incubator nearby. Johan walked around the room, searching for a thermometer. "One thing about a DIY lab is that you leave something somewhere, and it always ends up somewhere else," he told me.

I had joined Sosa at BioCurious so I could learn more about what CRISPR means for DIY scientists, and also to do a (hopefully) more successful experiment, with his help. We decided on a very basic goal: we would use the powerful editing tool to cut DNA he had already extracted from yeast cells. This task is easier than the one I tried in my kitchen, because you don't need to get CRISPR inside cells in order to slice the DNA. Professional scientists might use such a method as an intermediate step, such as when they need to cut and paste DNA together to make a gene as part of a bigger research project. "That's a type of experiment everyone does every day," explains Charles Gersbach, a professor of biomedical engineering at Duke University-though he notes that traditionally, researchers have used a type of protein called a restriction enzyme, not CRISPR, to do this.

Sosa and I pulled on latex gloves and carefully pipetted liquids into tubes to make our guide RNA from scratch-we first synthesized DNA strands with a specific sequence we wanted, used that as a template for the RNA, then destroyed the DNA and isolated the guide RNA from our test tube mixture. Later, we put the RNA in a new test tube, along with the other materials needed to make CRISPR work in this experiment: protein buffer, bovine serum albumin (a protein isolated from cows),water. Sosa sucked up the yeast DNA in his pipette. Without warning, the plastic needle-nose tip fell off into the tube of DNA. Someone, he told me, had donated the tips to their lab, but they weren't the correct size. "I guess you got the full DIY experience," he smiled, and pushed the plastic tip back onto the pipette. Then he picked up the Cas9 protein. "Here's the world-famous Cas9," he said, and handed it to me. I added it to our test tube.

Until CRISPR, DIY scientists didn't have an easy, cheap or reliable way to precisely edit DNA. Many of them couldn't afford the pricey and imperfect tools that professional scientists used for gene editing at the time. "Before CRISPR, there was TALENS [transcription activator-like effector nucleases] and zinc finger nucleases-older technologies that were not as precise or reliable," explains Sosa. "They were out of the budget and the time constraints of DIY scientists." Sosa says that if a DIY-er used those other technologies, it might cost him or her thousands of dollars to do a genetic engineering experiment. But with CRISPR, it's vastly more affordable, especially if you want to attempt an experiment more than once. "With TALENS, you try it once and fail," says Sosa. "With CRISPR, you can try it multiple times. That alone is a big deal."

This means that CRISPR gives DIY-ers a whole new way to do science. So far, Sosa and his lab mates have tried out CRISPR in a number of ways: cutting yeast genomes, slicing DNA inside E. coli cells, and attempting to modify the CRISPR system by shrinking it or attaching other molecules to it. Sosa has goals for his CRISPR research. "I want to understand how a cell really functions, and what are all the little things that happen in it," he explains. "And when something goes wrong [such as in diseases], how to fix it or make it do what I want."

After several hours, Sosa and I checked to see if CRISPR had cut our yeast DNA. We dyed our DNA-CRISPR mixture blue and ran it through an electrically charged gel, which separates bigger DNA pieces from smaller ones. Tiny channels in the gel run from one charged end to the other, and the sliced DNA strands are pulled through them towards the positively charged side. If our experiment succeeded, we should see two blue bands for the short CRISPR-cut DNA strands in one spot, and one blue band for a longer, uncut piece of DNA (our control) in another location.

Sosa carried the gel into the bathroom, where we turned off the lights and looked at it under blue light. I held my breath while I inspected the gel for markings. One light-blue band gleamed in the dark-the control-and another single band lit up the spot where we should have seen our CRISPR'd DNA. "I don't know what happened, but it doesn't look right," Sosa said, "I don't think it worked."

I left the lab feeling defeated, and headed back to San Francisco. Sosa texted me a few minutes later.

Hey, I figured out what happened. There was no DNA to start with, he wrote.

What happened? I texted back.

I think the DNA had either degraded or gotten too diluted, he wrote.

Even if we had got all the other parts (RNA, proteins, etc.) working, it didn't matter. We hadn't given CRISPR any DNA to cut. My second attempt at CRISPR had utterly failed.

BETTER, FASTER, CHEAPER

My own frustrating struggles with CRISPR aside, I wanted to see what professional biologists are doing with CRISPR, so I visited the lab of Nipam Patel at the University of California, Berkeley. After a quick tour of the lab, I sat down and stared into the microscope at a small, writhing marine creature: Parhyale hawaiensis, commonly called a beach hopper. At one centimeter long, Parhyale looks puny-you'd step on it at the beach without even noticing. But under the microscope, this female hopper resembled a giant translucent shrimp with many powerful, kicking legs. Parhyale is the star of this lab. "We're looking at how you develop an individual body," explains Erin Jarvis, a PhD student in Patel's lab, "And also how you build a body form over evolutionary time." And they're using CRISPR to do it.

With CRISPR, these researchers knock out so-called Hox genes in Parhyale. Hox genes are found in all animals, including humans, and they control the development of their body plans. Among other things, they determine what appendages-such as swimming legs, claws and antennae-grown on which section of the body.Knock out a certain Hox gene with CRISPR and Parhyale will grow forward-walking legs where it should have jumping legs, for example.

Parhyale has nine Hox genes, and Patel's team has knocked out seven of them. The researchers also have plans to add completely new genes to Parhyale using CRISPR-they've already done it once, by inserting a gene that codes for green fluorescent proteins, which allowed the researchers to visualize where a specific Hox gene is expressed in Parhyale."From an evolutionary perspective, this [gives] us insight into how body plans evolve between species," when comparing Parhyale to, for example, the well-studied fruit fly, Drosophila, explains Patel. "We believe that such evolutionary patterns help us understand the general mechanisms by which evolution creates animal diversity.... What we learn improves our knowledge about the function of these genes in other animals, including humans."

CRISPR has transformed how Patel and his colleagues do their research. His lab has looked at Parhyale for about 20 yearsnow. Before CRISPR they used another technique to knock out genes that required a lot more money, and even then, it wasn't very efficient. It cost them about $900 to knock out a single genein a group of Parhyale embryoswith their other method. The technique, called "RNA interference," silences expression of a gene-it doesn't genetically knock it out as CRISPR does. The problem was, sometimes the method didn't work at all.

Now it costs them less than $100 to knock out a gene. "Suddenly, with CRISPR, you don't have to decide, 'Which one gene do I want to put all my resources into?'" Jarvis says, "You can try a lot of different genes." And with CRISPR, they're able to break genes in up to 75 percent of Parhyale embryos, versus a maximum 25 percent success rate with the old technique. Even better, they now have the ability to mutate several genes at once with CRISPR, which means they can now see how genes interact. When the researchers had tried to mutate multiple genes with their old technique, it rarely worked. (Though Patel notes that the older RNA technique is still very useful for certain applications).

Their research takes less time with CRISPR, too-in a study Patel's lab published in Current Biology in 2015, they knocked out six Hox genes in about a year. Before that, they had already spent yearstrying to break a specific Hox gene with their old method, but were never able to do it. "Everything just goes faster," says Patel, a professor of genetics, genomics and development. "CRISPR-Cas9 is an incredibly elegant system, and it's very easy to control." It also makes it simpler to study more exotic creatures (beyond the standard flies and mice), such as animals like Parhyale or butterflies. "It's always been hard to work with a new organism," says Jarvis, "CRISPR is awesome because suddenly, you don't have to spend decades developing a model." As long as you have the sequence of the gene you want to target, you're set.

Patel's lab is hardly the only one capitalizing on CRISPR-scientists around the world are exploring all sorts of different uses for the gene editing tool, like wiping out malaria-spreading mosquitoes, finding new ways to treat cancer, or engineering disease-resistant crops. In July, researchers announced they had successfully edited the genome of viable human embryos with CRISPR; the technique allowed them to fix a disease-causing mutation in the embryos' DNA (though some are now skeptical of the researchers' results). Just a few weeks later, scientists in Massachusetts reported they had made a significant advance towards pig-to-human organ transplants. They used CRISPR to inactivate 25 viruses intrinsic to pigs' genomes, overcoming a big obstacle in making porcine transplants safe for humans.

PLAYING GOD

I had reached the end of my CRISPR experiment-so what had I learned? First, I found out it was not completely crazy for my roommate to wonder whether a DIY CRISPR kit in our fridge would make us sick. This year, German authorities restricted imports of the Odin DIY CRISPR bacteria kit after the Bavarian Health and Food Safety Authority tested two kits and found them to contain potentially pathogenic bacteria. But even theEuropean Center for Disease Prevention and Control concluded that there was little to worry about-that "the risk of infection by the contaminating strains in the kit is low for the users ... assuming that they are healthy people." (Zayner declined to comment on the record about the incident, but he publicly posted a response on Twitter, where he criticized the methodology used by the Bavarian agency and denied wrongdoing by his company. The kits are still available for purchase online through The Odin.)

As for my bigger question-could untrained DIY-ers actually achieve scientific breakthroughs?-I asked academic researchers what they thought. Dana Carroll, for his part, believes amateurs could make meaningful discoveries. "In the professional science community, people keep coming up with new ways to use this technology-people are really only limited by their imagination," he explains. "It's possible that people working in their garages or their kitchens will come up with a novel application or a solution to a problem that professionals just haven't gotten around to." And Carroll says it would be easy for a DIY-er to share any discoveries with researchers, by attending their talks or simply by contacting them through their Web sites. Yet he notes that the DIY community faces limitations, because amateur scientists likely would lack the necessary resources. "It's unlikely they will bring a major application all the way to fruition," he says, "But they could certainly get started on something."

Finally, what about the nightmare scenario: Is CRISPR so easy to use that we need to worry about biohackers-either accidentally or intentionally-creating dangerous pathogens? Carroll and others think that the danger of putting CRISPR in the hands of the average person is relatively low. "People have imagined scenarios where scientists could use CRISPR to generate a virulent pathogen, " he says. "How big is the risk? It's not zero, but it's fairly small." Gersbach agrees. "Right now, it's difficult to imagine how it'd be dangerous in a real way," he explains, "If you want to do harm, there are much easier and simpler ways than using this highly sophisticated genetic editing technique."

Back in Patel's lab, Jarvis replaced the squirming female beach hopper under my microscope with a tiny Parhyale embryo. Jarvis told me she knocked out a Hox gene called Abd-B in this one-the embryo will grow jumping legs where it should have swimming legs, and forward walking legs instead of anchor legs. At this point, it just looked like an opaque ball of goo to me.

Next to me, another grad student examined a fragment of a brown and goldbutterfly wing-Patel's lab is also knocking out butterfly genes with CRISPR to see how they build wing color. "An old grad student used to joke that we were genetically modifying the wings to make the Mona Lisa," Jarvis told me. I laughed and glanced back under the microscope. A puff of my breath suddenly struck the Parhyale embryo. It danced wildly around the petri dish, like a grain of sand caught in a windstorm.

# data[].attributes.attachment_link.title
Mail-Order CRISPR Kits Allow Absolutely Anyone to Hack DNA

# data[].attributes.attachment_link.description
"We aren't going to get sick, are we?" my roommate Brett asked me. He cringed as I knelt down and stuffed a plate of E. coli bacteria-which came as part of the DIY CRISPR-Cas9 kit I bought online-into our fridge next to cartons of eggs, strawberry jam, bottles of beer and a block of cheese.

# data[].attributes.body
Main idea:
Editing DNA via CRISPR
Arguments/facts:
- it has the ability to fix DNA errors
- It could create new treatments and cure diseases related to specific genetic errors
- It could also for example: create plants with larger fruits, make mosquitoes unable to transmit malaria, and reprogram drug-resistant cancer cells.
-

Arguments/facts/dialogue:
- It could be really useful.
- If it's that powerful it could also be a big risk
- It could neglect years and years of evolution just in a single process, which could also become dangerous for example plants in its habitat.
- CRISPR stands for clustered regularly interspaced short palindromic repeats and cas for CRISPR associated
-

Summary:
CRISPR is the fastest, easiest and cheapest of the gene-editing tools. CRISPR is actually a natural process but then edited to the liking of the scientist. As it was originally found as a bacterial immune system defending single-celled bacteria and archaea against invading viruses. The main idea is to cut and repaste a piece of DNA through the use of “molecular scissors” also known as the CAS9 protein. It can potentially even create treatments for genetic diseases and not just for humans, everything with a genetic code can be edited by CRISPR.

Andrea M. H.. [TED-ed], (2019, January 24). How CRISPR lets you edit DNA. Retrieved from https://youtu.be/6tw_JVz_IEc

# data[].attributes.attachment_link.title
How CRISPR lets you edit DNA - Andrea M. Henle

# data[].attributes.body
Main idea:
CRISPR is a powerful tool for editing genomes, meaning it allows researchers to easily alter DNA sequences and modify gene function. It has many potential applications, including correcting genetic defects, treating and preventing the spread of diseases, and improving the growth and resilience of crops. However, despite its promise, the technology also raises ethical concerns.

Arguments/facts:
- CRISPRs are specialized stretches of DNA, and the protein Cas9, which is an enzyme that acts like a pair of molecular scissors, capable of cutting strands of DNA.
- CRISPRs: The term "CRISPR" stands for "clusters of regularly interspaced short palindromic repeats" and describes a region of DNA made up of short, repeated sequences with so-called "spacers" sandwiched between each repeat.
- Scientists originally discovered the CRISPRs in bacteria in 1987, but they didn't initially understand the biological significance of the DNA sequences, and they didn't yet call them "CRISPRs".
- Scientists have been testing CRISPR to cure many diseases.
- Humans have been engineering life for thousands of years. We became very good at this, but never fully understood how it worked. Until we discovered the code of life, DNA. (A complex molecule that guides the growth, development, function, and reproduction of everything alive. Information is encoded in the structure of the molecule. Four nucleotides are paired and make up a code that carries instructions. If we change the instructions then the being carrying it also changes).

Dialogue:
- What would such uses mean for the status of people with existing genetic diseases if these are edited out of future generations?
- Should we make changes that could fundamentally affect future generations without having their consent?
- Could we trust us, humans, with this kind of technology?
- I think this is a revolutionary tool but I don't think we could trust humans with this.
- I'd say it's good that scientists decided to patent it and control its usage.

Summary:
CRISPR is an important tool for editing genetic information. With this tool, it became possible to edit genetic diseases, defects in DNA and viruses. The protein Cas9 is an enzyme that acts like a pair of scissors, which cuts strands of DNA and repasts a new modified strand. In other words, it is called “Gene editing” which is very recently discovered in 2017 in Japan. It was first discovered in the late 20th century and ever since the 21st century, this technology has been advancing at an exponential rate during these recent years. Although scientists are still testing it in therapeutic fields, they have already succeeded at germline (embryos) editing. This could be an important step in human history, but it is still doubted and debated as it is ethically concerning.

Aparna, H. Nicoletta, L. (2021, October 21). What is CRISPR? Retrieved from https://www.livescience.com/58790-crispr-explained.html

# data[].attributes.attachment_link.readable_text
CRISPR is a powerful tool for editing genomes, meaning it allows researchers to easily alter DNA sequences and modify gene function. It has many potential applications, including correcting genetic defects, treating and preventing the spread of diseases, and improving the growth and resilience of crops. However, despite its promise, the technology also raises ethical concerns.

In popular usage, "CRISPR" (pronounced "crisper") is shorthand for "CRISPR-Cas9." CRISPRs are specialized stretches of , and the protein Cas9 - where Cas stands for "CRISPR-associated" - is an that acts like a pair of molecular scissors, capable of cutting strands of DNA.

CRISPR technology was adapted from the natural defense mechanisms of and archaea, a domain of relatively simple single-celled microorganisms. These organisms use CRISPR-derived , a molecular cousin to DNA, and various Cas proteins to foil attacks by . To foil attacks, the organisms chop up the DNA of viruses and then stow bits of that DNA in their own genome, to be used as a weapon against the foreign invaders should those viruses attack again.

When the components of CRISPR are transferred into other, more complex, organisms, those components can then manipulate genes, a process called "gene editing." No one really knew what this process looked like until 2017, when a team of researchers led by Mikihiro Shibata of Kanazawa University in Japan and Hiroshi Nishimasu of the University of Tokyo showed, for the very first time, what it looks like when a CRISPR is in action, Live Science previously reported.

Related: Genetics by the numbers: 10 tantalizing tales

Key components of CRISPR

CRISPRs: The term "CRISPR" stands for "clusters of regularly interspaced short palindromic repeats" and describes a region of DNA made up of short, repeated sequences with so-called "spacers" sandwiched between each repeat.

When we talk about repeats in the genetic code, we're talking about the ordering of rungs within the spiral ladder of a DNA molecule. Each rung contains two chemical bases bound together: A base called adenine (A) links up to another called thymine (T), and the base guanine (G) pairs with cytosine (C).

In a CRISPR region, these bases appear in the same order several times, and in these repeated segments, they form what's known as "palindromic" sequences, according to the Max Planck Institute. A palindrome, like the word "racecar," reads the same forward as it does backward; similarly, in a palindromic sequence, bases on one side of the DNA ladder match those on the opposing side when you read them in opposite directions.

For example, a super simple palindromic sequence might look like this:

Short palindromic repeats appear throughout CRISPR regions of DNA, with each repeat bookended by "spacers." Bacteria swipe such spacers from viruses that have attacked them, meaning they incorporate a bit of viral DNA into their own genome. These spacers serve as a bank of memories, which enables the bacteria to recognize the viruses if they should ever attack again. You can also think of spacers like "Wanted" posters, providing a snapshot of the bad guys so they can be easily spotted and brought to justice.

Related: Going viral: 6 new findings about viruses

Rodolphe Barrangou and a team of researchers at Danisco, a food ingredients company, first demonstrated this process experimentally. In a 2007 paper published in the journal Science, the researchers used Streptococcus thermophilus bacteria, which are commonly found in yogurt and other dairy cultures, as their model, according to the Joint Genome Institute, part of the U.S. Department of Energy. They observed that after a viral attack, the bacteria incorporated new spacers into their CRISPR regions. Moreover, the DNA sequence of these spacers was identical to parts of the virus genome.

The team also manipulated the spacers by removing them and inserting new viral DNA sequences in their place. In this way, the researchers were able to alter the bacteria's resistance to an attack by a specific virus, confirming CRISPRs' role in regulating bacterial immunity.

CRISPR RNA (crRNA): CRISPR regions of DNA act as a kind of bank of viral memories; but for that stored information to be useful elsewhere in the cell, it must be copied, or "transcribed," into a different genetic molecule called RNA. Unlike DNA sequences, which remain lodged inside the DNA molecule, this CRISPR RNA (crRNA) can roam about the cell and team up with proteins - namely the molecular scissors that snip viruses to bits.

RNA also differs from DNA in that it's only one strand, rather than two, meaning it looks like just a half of a ladder. To build an RNA molecule, one part of the CRISPR acts as a template and proteins called polymerases swoop in to construct an RNA molecule that is "complementary" to that template, meaning the two strands' bases would fit together like puzzle pieces. For example, a G in the DNA molecule would get transcribed as a C in the RNA.

Each snippet of CRISPR RNA contains a copy of a repeat and a spacer from a CRISPR region of DNA, according to a 2014 review by Jennifer Doudna and Emmanuelle Charpentier, published in the journal Science. The crRNA interacts with the Cas9 protein and another kind of RNA, called "trans-activating crRNA" or tracrRNA, in order to help bacteria fend off viruses.

Cas9: The Cas9 protein is an enzyme that cuts foreign DNA. The protein binds to crRNA and tracrRNA, which together guide Cas9 to a target site on the virus's DNA strand where the protein will make its cut. The target DNA that the Cas9 will cut through is complementary to a 20-nucleotide stretch of the crRNA, where a "nucleotide" is a building block of DNA that contains one base.

Using two separate regions or "domains" on its structure, Cas9 cuts both strands of the DNA double helix, making what is known as a "double-stranded break," according to the 2014 Science article.

There is a built-in safety mechanism that ensures that Cas9 doesn't just cut just anywhere in a genome. Short DNA sequences known as "protospacer adjacent motifs," or PAMs, serve as tags and sit adjacent to the target DNA sequence. If the Cas9 complex doesn't see a PAM next to its target DNA sequence, it won't cut. This is one possible reason that Cas9 doesn't ever attack the CRISPR region in bacteria, according to a 2014 review published in Nature Biotechnology.

How CRISPR works as a genome-editing tool

Genomes encode a series of messages and instructions within their DNA sequences, and genome editing involves changing those sequences, thereby changing the messages they contain. This can be done by inserting a cut or break in the DNA and tricking a cell's natural DNA repair mechanisms into introducing the targeted changes. CRISPR-Cas9 provides a means to do so.

In 2012, two pivotal research papers were published in the journals Science and , describing how the bacterial CRISPR-Cas9 could be used to chop up any DNA, not just that of viruses. In this way, the natural CRISPR system could be transformed into a simple, programmable genome-editing tool.

To direct Cas9 to snip a specific region of DNA, scientists can simply change the sequence of the crRNA, which binds to a complementary sequence in the target DNA, the studies concluded.In the 2012 Science article, Martin Jinek and his colleagues further simplified the system by fusing crRNA and tracrRNA to create a single "guide RNA." Thus, genome editing requires only two components: a guide RNA and the Cas9 protein.

"Operationally, you design a stretch of 20 base pairs that match a gene that you want to edit," and from there, one can figure out what the complementary crRNA sequence would be, George Church, a professor of genetics at Harvard Medical School, told Live Science. Church emphasized the importance of making sure that the nucleotide sequence is found only in the target gene and nowhere else in the genome.

"Then the RNA plus the protein [Cas9] will cut - like a pair of scissors - the DNA at that site, and ideally nowhere else," Church explained. Once the DNA is cut, the cell's natural repair mechanisms kick in and work to piece the DNA back together, and at this point, edits can be made to the genome. There are two ways this can happen:

According to the Huntington's Outreach Project at Stanford University, one repair method involves gluing the two cuts back together. This method, known as "non-homologous end joining," tends to introduce errors where nucleotides are accidentally inserted or deleted, resulting in that could disrupt a gene.

In the second method, the break is fixed by filling in the gap with a sequence of nucleotides. In order to do so, the cell uses a short strand of DNA as a template. Scientists can supply the DNA template of their choosing, thereby writing-in any gene they want, or correcting a mutation.

Who discovered CRISPR?

Scientists originally discovered the CRISPRs in bacteria in 1987, but they didn't initially understand the biological significance of the DNA sequences, and they didn't yet call them "CRISPRs," according to Quanta Magazine. Yoshizumi Ishino and colleagues at Osaka University in Japan first found the characteristic nucleotide repeats and spacers in the gut microbe , and as the technology for genetic analysis improved in the 1990s, other researchers found CRISPRs in many other microbes.

Francisco Mojica, a scientist at the University of Alicante in Spain, was the first to describe the distinct characteristics of CRISPRs and found the sequences in 20 different microbes, according to a 2016 report in the journal Cell. At one point, he dubbed the sequences "short regularly spaced repeats" (SRSRs), but he later suggested that they be called CRISPRs instead. The term CRISPR first appeared in a 2002 report, published in the journal Molecular Microbiology and authored by Ruud Jansen of Utrecht University, with whom Mojica had been in correspondence.

In the following years, scientists also discovered Cas genes and the function of Cas enzymes, and they figured out that the spacers in CRISPRs came from invasive viruses, Quanta reported.

Among these pioneering researchers was Jennifer Doudna, a professor of biochemistry, biophysics and structural biology at the University of California, Berkeley, who went on to share the 2020 Nobel Prize in chemistry with Emmanuelle Charpentier, director of the Max Planck Unit for the Science of Pathogens. The two scientists are credited with adapting the bacterial CRISPR/Cas system into a handy gene-editing tool, Live Science previously reported.

Charpentier initially discovered tracrRNA while studying the bacteria Streptococcus pyogenes, which causes a range of diseases from tonsillitis to sepsis. Having uncovered tracrRNA as a previously unknown component of the CRISPR/Cas system, Charpentier began collaborating with Doudna to recreate that system in a test tube. In 2012, the team published their seminal work in the journal Science, announcing that they'd successfully simplified the molecular scissors into a gene-editing tool.

Some thought that biochemist Feng Zhang of the Broad Institute might also earn the Nobel for his own, separate work with the CRISPR system, Science Magazine reported. Zhang demonstrated that the CRISPR system works in mammalian cells, and based on this work, the Broad Institute earned the first patent for the use of CRISPR gene-editing technology in eukaryotes, or complex cells with nuclei to hold their DNA.

How has CRISPR been used?

(Image credit: KEITH CHAMBERS/SCIENCE PHOTO LIBRARY via Getty Images)

In 2013, researchers in the labs of Church and Zhang published the first reports describing the use of CRISPR-Cas9 to edit human cells in an experimental setting. Studies conducted in lab dish and animal models of human disease have demonstrated that the technology can effectively correct genetic defects. Examples of such diseases include cystic fibrosis, cataracts and Fanconi anemia, according to a 2016 review article published in the journal Nature Biotechnology. These studies have paved the way for therapeutic applications in humans.

In the realm of medicine, CRISPR has been tested in early-stage clinical trials as cancer therapy and as a treatment for an inherited disorder that causes blindness. It's also been investigated as a strategy for preventing the spread of Lyme disease and malaria from viral vectors to people, and it's also been studied in animal models of HIV as a way to rid infected cells of the virus, Live Science previously reported. One research team in China attempted to treat a human patient's HIV using CRISPR, and while the treatment wasn't successful in curing the infection, the gene therapy also didn't cause any harmful effects, Live Science reported.

"I think the public perception of CRISPR is very focused on the idea of using gene editing clinically to cure disease," said Neville Sanjana of the New York Genome Center and an assistant professor of biology, neuroscience and physiology at New York University. "This is no doubt an exciting possibility, but this is only one small piece."
Related: 10 amazing things scientists just did with CRISPR
CRISPR technology has also been applied in the food and agricultural industries to engineer probiotic cultures and to vaccinate industrial cultures (yogurt, for example) against viruses. It is also being used in crops to improve yield, drought tolerance and nutritional properties.

One other potential application is to create gene drives, a genetic engineering technique that increases the chances of a particular trait passing on from parent to offspring; this kind of genetic engineering derives from a natural phenomenon, where specific versions of genes are more likely to be inherited. Eventually, over the course of generations, the trait spreads through entire populations, according to the Wyss Institute. Gene drives could be used for various applications, such as eradicating invasive species or reversing pesticide and herbicide resistance in crops, according to a 2014 report published in the journal .

During the COVID-19 pandemic, the CRISPR-Cas9 system has been used to develop various diagnostic tests for the viral infection, BBC News reported.

In April 2017, a team of researchers released research in the journal Science that they had programmed a CRISPR molecule to find strains of viruses, such as Zika, in blood serum, urine and saliva.

On Aug. 2, 2017, scientists revealed in the journal Nature that they had removed a heart disease defect in an embryo successfully using CRISPR.

On Jan. 2, 2018, researchers announced that they may be able to stop fungi and other problems that threaten chocolate production using CRISPR to make the plants more resistant to disease.

On April 16, 2018, researchers upgraded CRISPR to edit thousands of genes at once, according to research published by the journal BioNews.

In addition, CRISPR has recently been used in the following ways:

However, despite its wide range of uses, the tool is not without its drawbacks.

"I think the biggest limitation of CRISPR is it is not a hundred percent efficient," Church told Live Science. That means, in a given experiment, CRISPR may successfully edit only a percentage of the targeted DNA. According to the 2014 Science article by Doudna and Charpentier, in a study conducted in rice, gene editing occurred in nearly 50% of the cells that received the Cas9-RNA complex. Meanwhile, other analyses have shown that depending on the target, editing efficiencies can reach as high as 80% or more.

The technology can also create "off-target effects" when DNA is cut at sites other than the intended target. This can lead to the introduction of unintended mutations. Furthermore, Church noted, even when the system cuts on target, there is a chance of not getting a precise edit. He called this "genome vandalism."

Potential risks and ethical concerns of using CRISPR

The many potential applications of CRISPR technology raise questions about the ethical merits and consequences of tampering with genomes. And in particular, a slew of ethical debates flared up in 2018 when He Jiankui, formerly a biophysicist at the Southern University of Science and Technology in Shenzhen, announced that his team had edited DNA in human embryos and thus created the world's first gene-edited babies.

He was subsequently sentenced to three years in prison and fined 3 million yuan ($560,000) for practicing medicine without a license, violating Chinese regulations on human-assisted reproductive technology and fabricating ethical review documents, Live Science previously reported. But even after his sentencing, He's experiments raised questions about how the use of CRISPR should be regulated going forward, especially given that the technology is still fairly new.

Illegal experimentation in human embryos represents an extreme misuse of CRISPR, of course, but even seemingly ethical uses of the technology could carry risks, scientists say.

In general, making genetic modifications to human embryos and reproductive cells such as sperm and eggs is known as germline editing. Since changes to these cells can be passed on to subsequent generations, using CRISPR technology to make germline edits has raised a number of ethical concerns.

Variable efficacy, off-target effects and imprecise edits all pose safety risks. In addition, there is much that is still unknown to the scientific community. In a 2015 article published in Science, David Baltimore and a group of scientists, ethicists and legal experts note that germline editing raises the possibility of unintended consequences for future generations "because there are limits to our knowledge of human genetics, gene-environment interactions, and the pathways of disease (including the interplay between one disease and other conditions or diseases in the same patient)."

In the 2014 Science article, Oye and colleagues point to the potential ecological impact of using gene drives. An introduced trait could spread beyond the target population to other organisms through crossbreeding. Gene drives could also reduce the genetic diversity of the target population, potentially hampering its ability to survive.

Other ethical concerns are more nuanced. Should we make changes that could fundamentally affect future generations without having their consent? What if the use of germline editing veers from being a therapeutic tool to an enhancement tool for various human characteristics?

To address these concerns, the National Academies of Sciences, Engineering and Medicine put together a comprehensive report with guidelines and recommendations for genome editing.

Although the National Academies urge caution in pursuing germline editing, they emphasize "caution does not mean prohibition." They recommend that germline editing be done only on genes that lead to serious diseases and only when there are no other reasonable treatment alternatives. Among other criteria, they stress the need to collect data on the health risks and benefits and to maintain continuous oversight during clinical trials. They also recommend that, after a trial concludes, trial organizers should follow up with the participants' families for multiple generations to see what changes persist in the genome over time.
This article includes additional reporting by Alina Bradford, Live Science contributor. Originally published on Live Science.
Additional resources

# data[].attributes.attachment_link.description
CRISPR is a powerful tool for editing genomes, meaning it allows researchers to easily alter DNA sequences and modify gene function. It has many potential applications, including correcting genetic defects, treating and preventing the spread of diseases, and improving the growth and resilience of crops. However, despite its promise, the technology also raises ethical concerns.