Dna Isolation Lab Report

PLASMID DNA ISOLATION, RESTRICTION ENZUME DISGESTION AND AGAROSE GEL ELECTRIPHORESIS
Abstract:
The gel is covered with an ion- containing buffer, such as (TAE), that controls the pH of the system and conducts electricity. overall DNA concentration was lower than expected. Using agarose gel electrophoresis is to separate and visualize the DNA fragment, which is produced by restriction enzymes .

Introduction:
The purpose of this experiment is to measure the size of the fragments of DNA and separate them after using the restriction enzyme. Dr. Jane Yeadon discovers the two kinds of plasmids that we used in DNA to GENOME laboratory, which are PJY5 and PJY48. She study two different fungal strains one of them is weak and another is strong, the
By using 0.8% of Agarose gel electrophoresis that about 30 ml to sit the wells. Carefully remove the comb from the gel without making bubbles that will gives a clear picture in the end.

Table 2 : the samples preparation:
Marker 1 2 3 4 5 6
2μl of DNA ladder + 10μl of water 2μl of
Dye+ 10μl of A 2μl of Dye+ 10μl of BamH1
A 2μl of Dye+ 10μl of Hpa1
A 2μl of Dye+ 10μl of B 2μl of Dye+ 10μl of BamH1
B 2μl of Dye+ 10μl of Hpa1
B

The purpose of adding Dye is to make DNA visible and enable to sink to the bottom of slots. The DNA samples + stain are loaded into the wells at one end of the gel. A power supply is added, where the positive electrode is at the opposite end to where the DNA has been loaded. Since DNA is negatively charged, it will move through the gel toward the positive end. Smaller fragments move easier and quicker, while larger fragments move slower.
When the power is shut off, the gel is photographed with a special UV camera that recognizes the dye. The smaller fragments will be at the bottom (faster moving), and the bigger on the top (slower moving)
We are able to measure the size of each band based on bp. It can be seen that, there are difference between lines A and B, the bands in B shows more light then the bands in A, therefore the concentration of DNA of B is higher than A. As a result, the intensity of B’s bands is greater than A’s bands. In line 2 the plasmid DNA is uncut which is expected while the line 3 A with Hpa1 plasmid DNA was uncut which is unexpected. This is due to different reasons such as the quality of enzyme was not good enough to cut the plasmid DNA, inaccuracies in the Elution step, the measurement of the restriction enzyme was not accurate, or there was any kind of contamination. Line4 has 2 bands one at the size of 6557 bp and the other at approximately 900bp which is lower and not that much clear. The band in line 5 Bam1 was 6800 bp which is expected and confirms our result that A is PJY5. In line 6 there is two bands on top of each other one is about 4361 bp and the other 4000 bp which means that both together is 8361 bp. This confirms our result that B is PJY5 (8361 bp). There are 3 types of DNA for our control: