Quavian Belton
Biochemistry I
Dr.Nicholas Panasik
October 6, 2015
PCR Optimization
Abstract
A Polymerase Chain Reaction (PCR) is a reaction that is set to catalyze the amplification of a copy of a piece of DNA. The aim of this lab was to optimize a PCR reaction by finding out the optimum concentration of MgCl2 needed. Six reactions were set up four with varying concentration MgCl2 and two primer control groups. The volumes and concentrations of each component of the PCR reaction was calculated and added up to 20uL as the ending volume. Each reagent was added to the tubes carefully. The tubes were placed in the PCR machine and ran on a 0.7% agarose gel. The samples were observed. Tube three had the biggest band that implied that it had the highest concentration in the MgCl2.
Introduction
PCR (polymerase chain reaction) is a technique enabling researchers to produce millions of copies of a specific DNA sequence in approximately 2 hours in a small test tube. This …show more content…
Once the concentration was found and the amount of everything reached 20uL , the process to pipette every reagent was added one by one with new pipette tips every time. Once filled the PCR tubes were placed in the PCR machine. First the tubes were ran at 95oC for 5 Minutes, followed by 30 seconds. Next 65oC for 30 seconds,72oC for 20 seconds, then back to 95oC for 30 seconds and steps followed 30 times.72oC for 7 minutes and finally hold the samples at 4oC. Samples were then run on a 1% agarose gel next to some DNA standards (Dr.Panasik.) 4 ul of sample dye was loaded to each tube and 10ul of each sample was loaded onto a 0.7% Agarose gel with Ethidium Bromide. The gel was electrophoresed for 1 hour and visualized under UV
Biochemistry I
Dr.Nicholas Panasik
October 6, 2015
PCR Optimization
Abstract
A Polymerase Chain Reaction (PCR) is a reaction that is set to catalyze the amplification of a copy of a piece of DNA. The aim of this lab was to optimize a PCR reaction by finding out the optimum concentration of MgCl2 needed. Six reactions were set up four with varying concentration MgCl2 and two primer control groups. The volumes and concentrations of each component of the PCR reaction was calculated and added up to 20uL as the ending volume. Each reagent was added to the tubes carefully. The tubes were placed in the PCR machine and ran on a 0.7% agarose gel. The samples were observed. Tube three had the biggest band that implied that it had the highest concentration in the MgCl2.
Introduction
PCR (polymerase chain reaction) is a technique enabling researchers to produce millions of copies of a specific DNA sequence in approximately 2 hours in a small test tube. This …show more content…
Once the concentration was found and the amount of everything reached 20uL , the process to pipette every reagent was added one by one with new pipette tips every time. Once filled the PCR tubes were placed in the PCR machine. First the tubes were ran at 95oC for 5 Minutes, followed by 30 seconds. Next 65oC for 30 seconds,72oC for 20 seconds, then back to 95oC for 30 seconds and steps followed 30 times.72oC for 7 minutes and finally hold the samples at 4oC. Samples were then run on a 1% agarose gel next to some DNA standards (Dr.Panasik.) 4 ul of sample dye was loaded to each tube and 10ul of each sample was loaded onto a 0.7% Agarose gel with Ethidium Bromide. The gel was electrophoresed for 1 hour and visualized under UV