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Polymerase chain reaction and gel electrophoresis
PCR - polymerase chain reaction
technique used to simplify small fragments of DNA
has revolutionised medical science
desired section of DNA is placed in a reaction chamber that has
various free nucleotide triphosphates
primers which allow replication from desired point
and a special heat stable version of DNA polymerase called Taq polymerase
was originally found in bacteria that live in hot springs
doesnt denature at the high temps used in PCR - can function in repeated cycles
DNA is heat enough to break hydrogen bonds that hold the 2 DNA strands together
called the denaturation phase
short primer sequence bonds to complementary sequences in DNA sample
known as annealing phase
extension phase - bonding of primers allows Taq polymerase to replicate DNA using the primer as a starting point
once DNA has been replicated, the strands are heated to point of separation & process begins again
each time a cycle occurs - amount of DNA doubles
if scientists needed to know more about an individual’s DNA, could use gel electrophoresis
identifies key features of DNA
uitilizes an electrical current to move molecules through a semisolid medium or gel
DNA molecules are separated by their size & amount of charge
DNA molecules hv negative electrical charge
to get fragments of appropriate size, DNA is digested using enzymes called restriction endonucleases
cut the backbone of DNA - create shorter segments
eukaryotic DNA is normally supercoiled by being tightly wound around histones to form nucleosomes
helps package DNA and allows it to fit better in nucleus
1st step: unwind coils to make strands accessible to enzymes
helicase then unwinds the double helix and separates the 2 DNA strands by breaking the hydrogen bonds
then, DNA polymerase moves along separate strands, uses them as templates, and builds a new strand of DNA by placing and attaching free nucleotides in a chain
prokaryotic DNA - naked DNA