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DNA 3 TEST 1 Notes 1

Lecture 1

What are the things needed for PCR? DNTPs(bases), Polymerase(for copying), Magnesium, Primers, DNA, Water, Thermal cycle, Buffer
What would you do to optimize a PCR Reaction? Add more resources. IE. DNTS, Polymerase, primers, Mg2 Run more cycles. Lower your annealing temperature.
In Amplification Define Stochastic Change. Using a very few cells, the Xsomes may not be represented equally, through transfer it could be lost. Establish a low limit for profiling. Ie 100 pg
MultiPlex PCR PCR allows for more than one region to be amplified to a greater degree than another.
Why do we add TAC at the end of a PCR cocktail? We add it last because it is activated by the presence of magnesium. We need it to interact with the primers not the nuculess or the dntps.
What does STR stand for? Short Tandom Repeat.
What are some of the advantages of STR markers? Small product sizes are compatible with degraded DNA Multiplex amplification with florescent detection enables high power of discrimination in a single test. Commercially available in an easy to use kit format. Uniform core STR loci = sharing of
What are the two main choices for STR Markers? Promega and Applied Biosystems (Life Technologies) FBI use Power Plex - Promega RCMP use Identifler/Profiler plus
What is BSA used for? BSA is used as a sponge to absorb inhibitors and stop them from interfering in a reaction.
What can you do to combat inhibition in a reaction. Use BSA to try and stop inhibitors. Or you can do a dilution. Diluting you sample also dilutes the inhibitor.
What is Low copy number? Low copy number is when you have less than 100 pico grams of template DNA for an amplification.
What were major events in DNA? Cult Massacre and the Romanov Family Murder
What types of extraction are there and give points about each? Kaiagen, Salt, Beads
What are the two main quantification methods? Gells for quality and Spectrophotometer for amount.
What are the denature,annealing, extension temperatures? Denature is 94-100, aneal is 55-60, extension is 72.
How many colours can a multiplex have? ?
What are STR markers>? They are an accordion like DNA sequence that occurs between genes.
Define the difference between a simplex and a multiplex? ?
What are the different type of nucleotide repeats? Di,Tri,Tetra,Penta,Hexa.
List values and disadvantages of STR kits. Advantage: Quality Control of materials, Saves time for end user, Improves consistency in results,Commen loci and conditions used(helps consitancy of databases),Easier for User. Disadvantages.Contents might not be known to user. High costs.
What are some major Forensic issues? Disucss points. Degraded DNA. PCR inhibition. Contamination. Copy Number.
Created by: Perrin Serroul Perrin Serroul