DNA manipulation Flashcards
Describe the denaturing phase of polymerase chain reaction
Denaturing - Heat DNA to 90oC to separate the DNA into single strands by breaking hydrogen bonds between complementary bases.
Describe the primer annealing phase of the polymerase chain reaction
DNA Primer Annealing - Cool to approximately 50oC to attach/anneal DNA primers to the single-stranded DNA.
Describe the elongation phase of the polymerase chain reaction
Elongation - Heated to approximately 72oC - DNA Taq polymerase copies the single DNA strands in a 3’ to 5’ direction using complementary base pairing.
State how many times the polymerase chain reaction is completed
35+ times
State the purpose of the polymerase chain reaction
To amplify the DNA so that there is enough to analyse
Name the fours stages of PCR
Denature/Dehydridise
Primers Anneal
Extension/Elongation
Repeat 35+
State the three temperatures of PCR
90 C
50 C
72 C
Outline the four stages of PCR
Denaturing - Heat DNA to approximately 90C to separate the DNA strands.
Primer annealing - Cool to attach approximately 50C to attach primers at the 3’ end of each DNA strand.
Extension (elongation) - Heat to 70C so Taq polymerase can copy strands in a 3’to 5’ direction.
Repeats - Process is repeated 35 times.
Reverse transcriptase is a
enzyme that copies mRNA into copy DNA
DNA primers are
Short single-stranded DNA fragments that attach to DNA to allow the binding of DNA polymerase
State the funciton of an endonuclease
Cut DNA at a specific recognition site
Describe how to create a DNA profile
Cut the DNA sample with an endonuclease
Place DNA sample in a well at the negative end of a gel
Turn on the electricity and the DNA fragments will move towards the positive end.
They will seperate based on size and charge.
State the purpose of standards in an agarose gel
These are DNA fragments of known size (bp) that can be used to estimate the size of the other fragments.
An endonuclease is
Bacterial enzyme that cuts DNA at a specific recognition sequence
Gel electrophoresis sorts DNA based on
size and charge