2 Polymerase Chain Reaction How is it used?Medical and biological research for a variety of applicationsDNA cloning, functional analysis of genes, diagnosis of hereditary diseases, genetic fingerprints (forensic science and paternity testing) and the detection of infectious diseases.
3 PCR Used to amplify (replicate) a single piece of DNA The original molecules are replicated by the DNA polymerase enzyme: doubles the number of DNA molecules (cycle)a cycle is repeated many times resulting in a “chain reaction”Makes it possible to amplify a piece of DNA: creating millions of copies of the original
4 Polymerase Chain Reaction A single segment is taken from a DNA sample to amplifyThe red segment is what is going to be replicated during PCR
5 Targeted DNAThe targeted DNA includes around 20 nucleotides, DNA primers, and the enzyme TAQ which are all used during PCR
6 Cycle 1The cycle begins with heating up the DNA mixture to 95° CelsiusThe heat denatures the DNA and breaks up the hydrogen bonds that hold the strands together .The temp. is then reduced to 60° so the primers can form hydrogen bonds
7 Cycle 1 cont. Temp. is raised to 72° Celsius At this temperature polymerization begins adding nucleotides to the DNA strandsThere are two double stranded copies of the DNA after 1 cycle
8 Cycle 2 Temp is raised to 95° again after the 2nd cycle there are 4 copies of the target DNA
9 Cycle 3 8 copies of target DNA are created after cycle 3 The more cycles, the greater the proportion of fragments with just target DNAHalf of the fragments consist of just target DNA and the other half also contains flanking DNA.
10 Cycle 4 & more 16 copies of the target DNA are created after cycle 4. With each cycle, the number of copies doubleThere are on average 25 cyclesOver 33 million copies of the target DN
11 PCR machines There are different types of machines All have a system that allows it to raise and lower the temperature of the DNA to allow PCR to occur
12 The PCR song!The PCR Song by Scientists for Better PCRThere was a time when to amplify DNA, You had to grow tons and tons of tiny cells. (Oooh) Then along came a guy named Dr. Kary Mullis, Said you can amplify in vitro just as well.Just mix your template with a buffer and some primers, Nucleotides and polymerases too. Denaturing, annealing, and extending, Well it’s amazing what heating and cooling and heating will do.[Chorus] PCR when you need to detect mutation (detect mutation) PCR when you need to recombine (recombine) PCR when you need to find out who the daddy is (who’s your daddy?) PCR when you need to solve a crime (solve a crime) [x2]