Agarose Gel Electrophoresis Lab Report

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3.2.5 Agarose gel electrophoresis The 1X TBE will be needed and to get this concentration, the dilution from the given 10X TBE buffer will be organized. In demand to prepared 500 ml of 1X TBE, a volume of 450 ml of 1X TBE and 50 ml of 10X TBE will be required. The 1.0% concentration of agarose gel will be prepared by mix the agarose powder and 1X TBE buffer. A 1.0g of agarose powder will be weighed and suspended in 100 ml of 1X TBE buffer. The mixture will be completely dissolved by put in the oven and will be let until the mixture are cool down a little bit. The GelRed stain will be added into the dissolved agar and the total volume of 100 μl of DNA stain will be added according to ratio which is the same as the volume of TBE. At 90V will…show more content…
Before centrifuged once again, the buffer PE will be added and the column will let to stand for 2 to 5 min. The flow-through will be discarded after centrifuged. The QIAquick column will be placed into a sterile microcentrifuge tube of 1.5 ml. A 50 μl of buffer EB will be added to the center of the QIAquick membrane to elute DNA and will be centrifuged for 1 min. A 30 μl of buffer EB will be added to the center of column to increase the concentration of DNA. The tube will let be stand for 1 min and will be centrifuged for 1 min. The elution buffer will be added and the incubation time up will be increased to 4 min. Therefore, it can increase the yield of purified DNA. At the end, the purified DNA will be analysed on the agarose gel. In ratio, a 1 volume of loading dye will be added to the 5 volumes of purified DNA. Before loaded to run the gel electrophoresis, the solution will be mixed by pipetting up and down. 3.2.7 Sequencing and analysis From the PCR purification from the previous step, the PCR product that received will be sent to the third party for a sequencing analysis. DNA laser gene software will be analysed the sequencing PCR product. In order to get homology sequence, the sequence will be explored by using the database from the GENBANK through BLAST
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