DNA Replication

What

-The synthesis of new strands of DNA, with the same base sequence as the original strand
-Yields 2 identical daughter molecules
-Semi conservative (due to complementary base pairing)

Needed for

-Growth
-Replacement
-Reproduction

Where

-In cells during the S-interphase of a cell cycle

Process

1.Helicase unwinds the double helical structure → results in a replication fork
2. Single stranded binding proteins (SSBP) prevet the strands from rejoining
3. Primase adds RNA primer provides a 3’ starting point for the DNA-polymerase
4.DNA polymerase adds free nucleotides in the 5’3’ direction, resulting in a lagging and leading strand
5. Complimentary base pairs are added (In the lagging strand, primers help add complimentary base pairs through which okazaki fragments are formed)
6.Ligase joins okazaki fragments
7.Daughter DNA molecules rewind to a double helix

PCR

polymerase chain reaction → copying of DNA artifically

PCR contains

-Tubes → eppendorfs, which contain the DNA sample, taq polymerase, primers and DNA nucleotides
-These are loaded into a thermocycler

Taq polymerase

-heat stable DNA polymerase
-enzyme from the thermus aquaticus → allows for high temps. to be used and speeds up the process

Cycle of PCR

1.Denaturation: DNA sample is heated → separates strands
2.Annealing → sample is cooled, allows primers to bind to both strands
3. Elongation → sample is heated to the optimal temperature for taq polymerase to function → copies strands and replicates them

-2x as much DNA is produced per cycle (doubled each time)

Gel electrophoresis

-Method of separating mixtures of positively or negatively charged macromolecules

Gel electrophoresis process

-Gel made of agarose
-positive and negative electrode on each end of the tank
-Charged macromolecules are attracted to one end
-DNA moves through the gel towards the positive electrode (due to phosphate)
-separates based on size of molecules + mass

Applications of gel electrophoresis

-DNA profiling
-Forensics
-Paternity tests