Flashcards in Lecture Two: Polymerase chain reaction and forensic DNA profiling (finished) Deck (114)
what was PCR first used for?
to identify skeletal remains using HLA DQα locus
At first to use PCR on VNTRs the sample had to be what?
By the 1990s PCR was used for what?
was used to amplify STRs
whats another word for
variable number tandem repeats
How many bp's do variable number tandem repeats have?
Amplified Fragment length Polymorphisms tandem repeats have an amplicon size smaller than what?
smaller than 1 kb to allow successful amplification
Human microsatellites are also known as what?
short tandem repeats
what stem typical core repeat for a STR?
whats the amplicon size of a STR?
Amplicon size 100-500 bp.
whats a Dinucleotide repeat?
same two repeat, example : (CA)(CA)(CA)
whats a simple repeat?
Tandem repeats with identical repeat units.
what do Non-consensus Alleles (microvariants) contain?
Contain incomplete repeats by one or more nucleotides
Consist of more than one type of simple repeat.
what does a Complex Repeat contain?
Contain several clusters of different tandem repeats with intervening sequences.
whats a simple definition for PCR?
Repeated copying of a selected region of a DNA molecule.
where is PCR done?
who won the noble prize for PCR?
Kary Mullis in 1985
what are the 3 main steps of PCR?
what happens at denaturation?
Denaturation at high
what happens at annealing?
Annealing at a cooler
annealing temperature (50°C-65°C)(~5°C below primers’ Tm; optimised experimentally)
what happens at extension ?
Extension at a temperature between the annealing and denaturing temperatures (usually at 72°C)
whats one cycle of PCR?
denature- heat to separate strands
annealing - hybridisation of primers
extension - DNA synthesis from primers
whats the average cycle length in PCR?
at what cycle is the correct size double stranded target created?
what is adenylation (A tailing)?
adding of a non-template nucleotide to the 3ʹ end.
what does incomplete adenylation show as?
this will appear as a split peak on the
electropherogram (epg) due to the different size PCR products.
what amplicon is favoured for adenylation?
In adenylation how do you go from -A amplicon to +A amlicon?
by a final incubation step.
what are the three phases of a PCR amplification curve?