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In the central dogma of biology, DNA replicates itself in order to do cell division. Then, it transcribes into RNA to travel to the organelles to make proteins, (It transcribes into RNA so that the genetic information can leave the nucleus) and then the RNA goes to its designated organelle, where its translated for the creation of proteins.

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In the replication process there are some key enzymes we need to know:

Topoisomerase: Untwists DNA molecule

Helicase: separates the 2 strands of DNA into lagging strand and leading strand

SSB: Stabilises the molecule so that it doesn't twist again

Primase: adds primers

DNA Polymerase III: adds nucleotides in 5' to 3' direction.

DNA Polymerase I: changes primers (RNA) to DNA

DNA Ligase: Bonds gaps (incomplete bonds that could occur by the Polymerase I)

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Process:

1. topoisomerase untwist the DNA and helices separates it into lagging and leading strand

2. The SSB molecules stabilise and hold the 2 strands in place so they don't join back together or twist.

3. (Then the rest of the enzymes start working to add another strand to each loose strand, and get 2 structures of DNA. These enzymes work in direction 5' to 3', so one strand (Leading strand) will work in a continuous manner because 5' end is behind, and it works forward towards 3' where the helicase and topoisomerase are working. The other strand (lagging strand) will have 5' where the 2 strands are being separated and 3' behind, so it will work in a discontinuous manner, where the enzymes will go forward and work their way back, and as soon as more strand of DNA gets separated, they will go back forward and work back again, working in fragments.)

4. the primase will add primers (fragments of RNA) to the strands to start the process of creating the new structure.

5. then the DNA polymerase III will attach to those primers and start working, creating the nucleotides (DNA) from the primers (RNA) in direction 5' to 3'.

6. Then the DNA polymerase I will take away the primers that were put (because the RNA isn't useful) and change it for DNA.

7. After that the ligase enzyme will check and close any possible gaps in the DNA.

8. The leading strand will work continuously forward with no problem but since the lagging strand will work backwards in fragments, there will be unjoined fragments of DNA that the ligase will bond. These unjoined fragments are called Ozaki fragments.

9. In the end, the 2 strands will have an opposite strand for each and we will have two structures of DNA composed of 2 strands each.

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Transcription is a process from which a strand of DNA is converted to RNA, so it can travel to its final location to create the codified proteins, hormones, etc. It is composed of 3 stages: Initiation, elongation and termination. The final result of this process is a detached strand that is a copy of the desired genetic information that will be able to travel around the cell (for a short period of time because RNA is reactive and will degrade over time).

3 STAGES:

Initiation: the desired geen is located, and the RNA Polymerase (only enzyme in transcription) attaches to a promotor (enzyme that locates beginning of the desired gene) and a termination signal locates the end of the desired gene.

Elongation: The RNA Polymerase, after having located the gene between the promotor and termination signal, elongates the gene (in double helix structure) and opens it into a bubble, where there is the top and bottom string. The bottom string is the template strand, and the RNA polymerase creates a copy of that strand and its information, working in the direction 5' to 3' and detaches strand after getting to the end of the gene.

Termination: After elongation, the opened gene (bubble) closes back into double helix, and we are left with a detached strand that is a copy of the genetic information of the gene, that will undergo some final post-transcription modifications to get protected and be able to travel around the cell.

In transcription there is no thymine, so adenine (A) which used to combine with Thymine (T) now combines with a new pyrimidine called Uracyl (U) which is useful for protein synthesis, unlike thymine (T).

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In Eukaryotic cells, after transcription, a series of modifications take place in the mRNA (messenger RNA). The RNA is made up of introns and exons, inside of which, only the

exons contain usefull information. Due to this, a type of splicing takes place to remove the undesired introns.

2 types of splicing can take place:

a) Splicing; in which the exons are lined up accordingly. (E1, E2, E3...)

b) Alternative splicing; in which the exons are lined up randomly to create a different type of protein. (E3, E6, E1...)

In alternative splicing, there could be thousands of different combination used to make thousands of different proteins.

After the splicing, the mRNA needs to make a small trip inside the cell to the Endoplasmic Reticulum, in which, it needs a small protection to stabilise the strand. Here is where a 5´ (in 5' end) cap of guanine is placed on one end, and a series of 3´ Adenine that acts as the tail (in 3`end).

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For translation there are some definitions we need to know:

mRNA- messenger RNA, product of transcription that contains "instructions" for a protein.

tRNA- transfer RNA, transports an amino acid

Amino Acid (aa)- monomers of a protein. A chain of aa is a protein.

Ribosome- organelle where protein synthesis takes place.

Codon- Group of 3 nucleotides that codifies for 1 aa.

Anticodon- at the bottom of tRNA, attaches to a codon of mRNA inside the ribosome.

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The main components of the process of translation are the mRNA, tRNA, amino acids and Ribosomes.

The order is:

1. The first step in translation comes when a tRNA attaches itself to the mRNA strand in the ribosome. More specifically, the anticodon of the tRNA attaches to a codon of the mRNA. This now passes throught the big-subunit of the ribosome into the "A" part.

2. This next step comes when the strand passes one step to the left, and a new tRNA attaches to the next codon of the mRNA. Here the amino acids of the tRNAs inside the ribosome create a bond and connect to each other.

3. One more step to the left, and another tRNA attaches itself to the next codon of the mRNA while the previous tRNA and its amino acids joins with the recent one. Simultanously, the first tRNA leaves the ribosome by detaching from the amino acids that are creating a chain of proteins. The tRNA enzyme is then reusued for next processes, when they will attach to a loose amino acid and redo the process.

The process ends whenever the codon has a specific code for termination, which makes the amino acid chain (protein) leave for its next functions. The big subunit is the space inside the ribosome and the 3 spots the tRNA passes through is the "A" part when it enters and attaches to a codon of the mRNA, the "P" part when it moves to the middle section and the amino acid creates a bond with the new amino acid that has entered the ribosome via the new tRNA that has attached to the previously passed "A" part (the tRNA is attached to the next codon). Then the tRNA goes to the "E" part where the tRNA and amino acid detach, the tRNA leaves to be reused and the amino acid is held together by a forming chain of amino acids that will later become a protein, hormone etc. Every codon attatches to a specific anticodon of a tRNA that has a specific amino acid.

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Mutations are mistakes during the processes of DNA replication or cell division. A new sequence of DNA is produced.

Mutagens are agents that facilitate mutations to occur. we are studying 3 types:

1. physical mutagens (like x-rays or UV light)

2. Chemical mutagens (like pollutants, tobacco, etc.)

3. Biological (some viruses can produce mutations)

Mutations on different processes of DNA replication will have more or less impact, depending on the stage the mistake occurs in.

There are also types of Mutations

Base Substitution

Missense Mutations: Change of base changes the amino acid inserted in the protein.

Nonesense Mutations: Change in the base causes termination of protein being produced.

Silent Mutation: Change in base causes no change in protein because the new base codifies for the same protein.

Insertion

This occurs when an extra nucleotide is added to the DNA strand during replication. This can happen when the new strand (in DNA replication) being produced "slips" or "wrinkles" and allows the incorporation of an extra nucleotide.

Deletion

Occurs when a "wrinkle" forms in the DNA template strand and causes a nucleotide to be omitted from the replicated strand.

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Genetic engineering is the modifying of an organism at genetic level in order to do something it wasn't able to do.

PCR's a Basic Staple

The basis of a PCR is to make exponential copies of a sequence of DNA. Specifically, primers will be used to find specific sequences.

PCR= Polymerase chain reaction

Electrophoresis

Is putting the PCR sample on a gel and running electricity through the gel. This causes the genes to move (smaller genes go furthest than larger genes). This separates the genes (fragments of DNA) so we can see them clearly (It is like a code). After we do the electrophoresis, the results are compared with the same sequence of DNA of other people (that are/aren't infected with a virus for example) to see the results and if the sample person coincides with a gene that the infected person has and the healthy person doesn't, we can deduce if the sample persons infected or not. (This was used greatly during COVID-19 pandemic).

In another case, the genes are added coloured nucleotides, passed through electrophoresis and the results are shone on a laser to see the coloured nucleotides attatched to the genes. This helps us find information because as the genes are separated from largest to smallest through electrophoresis, we see the coloured nucleotide that is Adenine for example, and we know that adenine pairs with thymine, so we know that the nucleotide next to the coloured gene has thymine. And as we go further, we find the next nucleotide because 1 nucleotide less is smaller and goes further, so in the end you can know all the sequence of DNA.

Plasmids and their Manipulation

Plasmids are small extrachromosomal DNA molecules that are loose in bacteria. In genetic engineering, these plasmids are taken and cut out a section to then put in a desired gene in the bacteria to produce Insulin for example. Also, an antibiotic resistance gene is added to the bacteria so that the gene produces the desired product and is resistant. This bacteria then without knowing will produce insulin, (in this case) and as it doesn't need it, it will expel its insulin to its surroundings. In the case that this genetic modification doesn't work, we add antibiotics to kill the bacteria that didn't work, and as the bacteria that worked have an antibiotic resistance gene, they won't die. In the end you will have a tank of antibiotic resistant bacteria that produce insulin. (In this specific case).

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As we see in the image, we have a plasmid that is cut with a restriction enzyme in the restriction site, and the desired gene is added (in this case called inserted gene) along with an antibiotic resistant gene so that the plasmid is resistant and productive. Like copy of a gene pasted on the plasmid.