User:Elaine Marie Robbins/Notebook/CHEM-496/2011/09/21

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Gel Electrophoresis of PCR Product Main project page
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Objective

To determine whether or not the PCR performed on 09/20/11 of the DNA coding for a mutant GFP was successful using agarose gel electrophoresis.

Description

  1. Make a 1% agarose gel by adding 0.25 g of 0.01 g/mL of agarose to 25 mL of 1x tris base, acetic acid, and ethylenediaminetetraacetic acid (TAE) buffer and heating the mixture in the microwave for 40 s.
  2. Mix 1 μL of 6x gel loading dye with 5 μL of the DNA sample.
  3. Prepare a ladder containing DNA, glycerol, and gel loading dye.
  4. Load DNA solutions and run the gel at 80 V for 40 min.
  5. Remove gel and place in TAE and ethidium bromide (EtBr) solution, mix for 15 min.
  6. Rinse gel with TAE for 5 min.

Data

A photo of the electrophoresis gel is displayed below:

image:DNA gel 092111 003.jpg

The first column is the ladder. The next columns are groups 1, 2, 3, 4, and 5.

Notes

Gel ladder: 500 bp, 1000 bp, 2000 bp, 3000 bp, 4000 bp, 5000 bp, 6000 bp, 7000 bp, 8000 bp, 9000 bp, 10000 bp

An optimum gel would have used 1.5% agarose.




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