Polymerase Chain Reaction Lab Report

CHROMOSOME 16: PV92 PCR INFORMATICS

AIM: To apply PCR to amplify a specific sequence of a DNA.

INTRODUCTION:

PCR (POLYMERASE CHAIN REACTION):

Polymerase Chain Reaction, shortly known as PCR, is an easily understandable and cheap method that is used to make many copies of a specific section of DNA over and over again. It is one of the revolutionary methods found in the field of Biotechnology. PCR uses the DNA polymerase which is an enzyme that synthesizes a complementary new strand to the pre-existing template strand of DNA. The DNA polymerase used is called Taq Polymerase. In this case, the template DNA is the segment of DNA which is wanted to be copied. Primers are needed in order to add a nucleotide, the component which is structural for

Then the saline was expelled back into the cup. 1,000μl of the oral rinse was transferred into the 1.5μl micro centrifuge tube. The tube was then spanned in a balanced centrifuge for 5 mins in order to make a pellet of whitish cells at the bottom of the tube. The supernatant was discarded and the pellet was re suspended by the process of vortexing. After this process 20μl of the re suspended cells were added into a tube containing Instagene Matrix. The need of using Instagene is due to making the process of preparation of DNA samples quicker and easier. By using Instagene the cost is less and it supplies a faster PCR quality template of DNA (8). Then the tubes were vortexed again in order for the contents to mix properly and was incubated for 15 mins in 56 degree water bath. Incubation is used in order to split up the tissue which holds the cells and stops the enzymes from working. A week later the tubes that contained the DNA were taken from the refrigerator and centrifuged for 2 mins. Then 20 μl of the DNA supernatant was added to the PCR tube and 20 μl of Master Mix was added to this tube also. Master Mix contains DNA enzymes, many salts, only magnesium, nucleotides and a reaction buffer in order to simplify PCR. Then the PCR tube was placed in the thermal cycler for running PCRs (10, 11). After all these steps PCR samples were added 10 μl of Orange G loading dye and the samples were added to the wells of the gel one by one. 1st lane contained MMR, 2nd lane contained Homozygous (+/+) control, 3rd lane contained Homozygous (-/-) the 4th lane contained Heterozygous (+/-) and the remaining last 6 wells contained the student samples. The power supply was turned on and the samples were electrophoresed for 30 mins. Then they were