Dna Purification Lab Report

The purpose of module E is to learn several DNA techniques in the lab including DNA purification with solubility and absorption, plasmid transfection of E.coli, colony screening by PCR and quantitative PCR.
First part of the experiment E1 show the purification method of DNA through solubility. E. coli lysate mixed organic solvents to purify the DNA present in solution. First, the lysate was mixed with phenol/chloroform, then vortexed, and centrifuged. We extracted the aqueous layer and combined with sodium acetate and cold ethanol and then centrifuged and filtered. The ethanol causes the DNA to precipitate since the contaminants were removed from the previous precipitation. The remaining pellet was virtually invisible, but, when re-suspended with TE buffer, produced a reading in the UV spec.
These plasmids can be inserted into the bacterium’s genome which may increase their chances of survival through diversity. This method was done in the lab by using vector transfection. Vectors are the plasmids that are inserted into bacterium for protein expression. Vectors are added to solution with bacteria exposed to heat-shock so they take up the DNA. These bacteria are then allowed to grow and replicate on agar included vector. Some indicator is also added the vectors to indicate which colonies have inserted the vectors.
Experiments E2 and E3 were transfection of vectors and analysis of transfection respectively. The vector being used included an ampicillin gene as its selective gene and a fluorescence gene induced with arabinose. Three agar plates were prepared with three different samples. Sample 1 was a control and included only LB, sample 2 included LB and ampicillin, sample 3 included LB, ampicillin, and