BME100 f2015:Group5 1030amL4
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LAB 4 WRITE-UP
Research and Development
PCR - The Underlying Technology
Template DNA is a strand of DNA contains the targeted sequence of DNA that we are going to isolate with primers and mass replicate through PCR. Primers are single-stranded DNA sequences that complement the starting codon sequence. Primers are highly specific, targeting a specific sequence of DNA. DNA polymerase II begins the replication process of the target sequence starting from the end of the primer once it is bond to its complementary sequence. Taq polymerase is a DNA polymerase II enzyme taken from Thermus aquaticus - an organism that lives in extremely high temperatures such as in hot springs or vents. Due to its environment, the DNA of T. aquaticus can withstand extreme temperatures without denaturing, unlike human DNA. This characteristic of T. aquaticus DNA allows us to use enzymes from said organism to replicate a target sequence of DNA via thermoshock without destroying the enzymes needed to replicate the sequence.
The general term used to describe the four deoxyribonucleotides - dCTP, dGTP, dATP, and dTTP - is 'dNTPs.'
Base-pairing occurs during the annealing and the extending phases. During the annealing phase, the nucleotide sequence of the primer must successful pair with the correct nucleotide sequence on the single-stranded DNA. During the extending phase, Taq polymerase adds complementing nucleotides to the target sequence, replicating and extending it.
SNP Information & Primer Design
Background: About the Disease SNP
Nucleotides are organic molecules that are the "building blocks" of DNA. These "building blocks" are known more specifically as adenine, cytosine, thymine, and guanine. When two or more clearly different phenotypes exist in the same population of a species - we call it a polymorphism. When referencing the term "Disease SNP," we associate it with a single change in one of the bases, in turn causing the creation of undesired proteins and possibly leading to serious diseases.
Our SNP variation is found in Homo Sapiens on chromosome 16. This pathogenic SNP is known to be directly associated with the MC1R gene - a marker for melanoma. MC1R - or Melanocortin 1 receptor - functions as an encoder for a receptor protein for the alpha melanocyte stimulating hormone receptor. For this specific disease, the disease-associated allele contains the TGG sequence.
Numerical Position: 89919736 (Numerical Position 200 bases to the right would be 89919936)
Primer Design and Testing
For the non-disease forward primer and the non-disease reverse primer, our tested and validated results were chromosome 16 and 220 base pairs (B.P.). As for the disease specific primers, the tested and validated results got no matches.