D1.1: DNA replication

DNA replication use complimentry base pairing in order to create a new copied strand which creates exact copies of DNA with identical base sequences

DNA replication is essential for cell division during growth and repair of damaged tissues, while it also ensures that each of the new cells receives its own copy of the DNA.

DNA replication is said to be semiconservative because each newly made DNA molecule has one original and one new strand of DNA.

As DNA replication is semi-conservative (one old strand an d one new strand make up the new DNA molecules), this complementary base pairing allows the two DNA molecules to be identical to each other as they have the same base sequence. The new strands formed are complementary to their template strands but also identical to the other template. Therefore, complementary base pairing has a big role in the conservation of the base sequence of DNA.

so each original strand can act as a template for building a new, complementary strand as it exposes the nucleotide bases, enabling enzymes like DNA polymerase to match free nucleotides to their partners (A-T, G-C) and synthesize two identical DNA double helices from one.

DNA helicases are essential during DNA replication because they separate double-stranded DNA by breaking the hydrogen bonds between complementary base pairs to form single strands allowing each strand to be copied.

DNA Polymerases are a group of enzymes that catalyse the synthesis of DNA during replication. They do this by adding complementary nucleotides to a template, working in the 5' to 3' direction and requiring a primer to start

DNA polymerase III: This enzyme synthesizes the leading strand continuously and also extends the Okazaki fragments (short, discontinuous pieces of new DNA synthesized on the lagging strand during DNA replication) on the lagging strand (the new DNA strand).

DNA polymerase I: This enzyme removes the RNA primers and fills in the gaps with DNA on the lagging strand.

Polymerase Chain Reaction

Allows rapid amplification of a specific segment of DNA

Denaturation: The DNA is heated to about 95°C for 30 seconds to separate the hydrogen bonds in the double-stranded DNA into two single strands.

Annealing: The temperature is lowered to about 54°C for 30 to 60 seconds, allowing short RNA primers to bind to the specific regions flanking the target DNA sequence.

Extension: The temperature is raised to about 72°C, and Taq polymerase (a heat-resistant enzyme) adds nucleotides to extend the DNA strand, synthesizing the new DNA strand.

Ends with two double stranded DNA

the number of bands directly indicates the number of different-sized DNA fragments in a sample, while their relative sizes are deduced by their distance from the well: smaller fragments travel faster and further down the gel, while larger fragments move slower and stay closer to the well, compared to a DNA ladder (marker) of known sizes.

STEP 1: Prepare samples. - DNA is cut into varying sized pieces with restriction enzymes
STEP 2: All pieces are placed into small wells in the agar (in a buffer solution)
STEP 3: A current is applied to the gel. DNA has a negative charge
STEP 4: The length the DNA moves is inversely proportional to the size of the fragment (smaller move farther)
STEP 5: Visualize and document bands. - DNA is stained and banding pattern is observed

Each phosphate group in DNA carries a negative charge, which means that DNA can be pushed by an electric voltage, from the negative electrode toward the positive electrode.
This process separates the DNA fragments by length because shorter fragments of DNA move more quickly through the gel, and longer fragments move more slowly.
At the end, each DNA sample produces a unique banding pattern, so we can compare DNA samples.
If the banding pattern of two lanes match, then those two DNA samples came from the same source.

- Forensic sciences
- Paternity testing
- DNA profiling

1) Swab tak from noise/throat wher virus particles and rna is rinsed in saline to produce a liquid sample
2)RNA is converted to DNA using enzyme reverse transcriptase
3) Same is placed in PCR to amplify specfic base sequences (markers for straubs of COVID) 35 cycles are sued
4) In PCR fluorescent markers are attacted to DNA produced.
5) fluorescent markers are montinered and if they are rise above a certain level its positive for COVID

advantages:
sensitive - 1 molecule of RNA amplified to produce 35 billion molecules of DNA
Very specific - Primers can be designed so only 1 strain can be detected

Disadvantages
material and equipment is expansive and PCR is only aviable in testing labs

DNA prolfing distingiushes indivuals using sequences known as short tandem repeats
thiirteen or more tandem repeats are widley used in DNA profiking because its very unlinkey that two humans would have the same number of repeats of each of these short tandem repeats

1)sample of DNA obtained
2)selected tandem repeats are copied by PCR - at least 13
3)DNA is produced by PCR is sepereted according to lengeth - thereofre number of repating using gel electrophoresis
4)Patterns of bands of DNA produced on gel (these bands are always the same with DNA taken from one individual)

- Forensic sciences
- Paternity testing

Using the gel electrophoresis look for if evidence is at the same level of a suspect the closer the evidence is to the suspect the more related the suepect is to the evidence

- Blood
- Semen
- Saliva
- Urine
- Feces
- Hair
- Teeth
- Bone
- Tissue
- Cells