Techniques used for Profiling procedures forensically
Restriction Fragment length Polymorphisms (RFLP); Short Tandem Repeat Multiplex (STR Multiplex); [Amplified Fragment Length Polymorphisms (AFLP) ]
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Which part of the DNA do you need to profile
Small areas (loci) of DNA which differ markedly among individual people
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What must the DNA sequence contain for Restriction Endonucleases (REs) to start cutting (cleaving)
sequence must be palindronic - appears on both strands at the same place on the DNA
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Where can you find Variable Number Tandem Repeats – VNTRs
In the Restriction Fragment Length Polymorphism (RFLP)
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How many repeats can yu find of Variable Number Tandem Repeats (VNTRs),
Each repeat typically contains 20 –100 base pairs, repeated from 1 to 30 times
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How do we determine the size of the restriction fragments?
The fragments are separated on the basis of size by Agarose Gel Elecrophoresis
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How does Agarose Gel Elecrophoresis work
Fragments are drawn through the gel by an electrical current; -ve charged DNA attracted to +ve terminal; Smaller fragments move further than bigger fragments
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how was Polymerase chain reaction (PCR) discovered
Heat resistant DNA polymerase isolated from a bacterium Thermus aquaticus
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What reaction does thermal cyclers cyclically change the temperature
DNA; DNA polymerase; source of nucleotides; multiple copies of 2 primers
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In theory, what is the purpose of thermal cyclers
Each thermal cycle doubles the quantity of targeted DNA
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What is the mixture inside the Polymerase chain reaction (PCR) tube
The DNA to be amplified; Taq polymerase; DNA primers Nucleotide sources (dNTPs) with all 4 bases (A,T,C,G)
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How does Polymerase chain reaction (PCR) work
mixture of components in tube; heated at 90 to denature DNA; cooled to 60 for primer annealing; heated to 72 for Taq polymerase to extend the primers- synthesis complimentary DNA strand
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how to get another cycle of Polymerase chain reaction (PCR)
after 2 exact copies are produced; Heat to 90 oC and the whole process begins again
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Why do DNA profiling use Short tandem repeats (STR)
abundant throughout the genome; Highly variable in populations; Small size means they are likely to remain in degraded DNA
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What is the problem with Short tandem repeats (STR)
less polymorphic than VNTRs, so you need more STR loci; 12 STR loci = 5 VNTR loci
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What kind of electrophoresis is used for Short tandem repeats (STR)
Poly Acrylamide Gel Electrophoresis (PAGE)
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Why do Short tandem repeats (STR) use Poly Acrylamide Gel Electrophoresis (PAGE)
Because STR fragments are short, gel with greater resistance to DNA movement must be used to separate them
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Why is it difficult to use Second generation multiplex (SGM)
because primers must share same annealing temp, not bind to each other or the wrong DNA loci
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What kind of DNA profiling does the FBI use
CODIS STR profile
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What is DNA 17
European standard for DNA profiling; Analyses 16 STR loci –spread over several chromosomes; gives size of 32 loci; Profile based on 19 or more locai has a discriminating power of 1 in 1 X109
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What is the problem with DNA 17 when conducting a crime scene
does not always yield a full profile; 4 STR loci gives a discriminating power of 1X 106; 4 loci the smallest profile entered on the British National DNA Database
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examples of human error in cases
Huston forensic lab in 2006;Meredith Kercher case; Malpractice at Randox Testing Services; UK forensic regulator report for 2016-17 – highlighted growing concerns about the failure of some forensic firms
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does DNA profiling work with everyone
Monozygote twins – exactly the same DNA; person who got a bone marrow transplant
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Why is mitochondria DNA useful
good for old and badly degraded tissues
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Other cards in this set
Card 2
Front
Techniques used for Profiling procedures forensically
Back
Restriction Fragment length Polymorphisms (RFLP); Short Tandem Repeat Multiplex (STR Multiplex); [Amplified Fragment Length Polymorphisms (AFLP) ]
Card 3
Front
Which part of the DNA do you need to profile
Back
Card 4
Front
What must the DNA sequence contain for Restriction Endonucleases (REs) to start cutting (cleaving)
Back
Card 5
Front
Where can you find Variable Number Tandem Repeats – VNTRs
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