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Process Of Dna Extraction Of Animal Tissue

1021 words - 5 pages

Extraction and purification of nucleic acid is the first step in most molecular biology studies and all in recombinant DNA techniques. DNA extraction is a process of isolation of Genomic DNA from RNA, protein, and lipid. Basically, a genome is the complete genetic contents in chromosomes of eukaryotes and prokaryotes or in the RNA and DNA of viruses. Genome is the total haploid set of organism’s genes (Campbell, Reece, Taylor, Simon, & Dickey, 2011). The purpose of DNA extraction is to obtain DNA in a relatively purified form which can be used for further investigations such as PCR and DNA sequencing. Many different methods are available for isolating genomic DNA. Modification and ...view middle of the document...

This is to make sure the cell membrane is completed lysis. On the other hands, if the tissues do not dissolved completely after two hours Proteinase K can be added more.
Furthermore, chloroform-isoamyl alcohol is added and shaked for 2-3 minutes. Then, it is centrifuge at 13000rpm for 10 minutes. Chloroform-isoamyl alcohol (CIA) also acts as detergent to bind to protein and lipids of the cell membrane of the tissue. Then, both protein and lipids are dissolved by the breakdown of membrane due to the disruption of bonds that hold the membrane together. In addition, the function of the CIA is to separate the contaminants into the organic phase and aqueous phase for a nucleic acid (Farshad, Craig, & Naderia, 2013). This is because the CIA is denser than the water solution. After centrifuge is completed, only the upper layer of the supernatant is taken and transfer to a new appendorf tube. In addition, only the upper layer of the supernatant contains the DNA.
The next step is DNA precipitation. Deoxyribonucleic acid (DNA) precipitation is the process of the separation of nucleic acid from contaminant (Herzer, 2001). In this step, 500µl of cold absolute ethanol (99%) is added and mixed well with the supernatant in the newly labeled eppendorf tube. Ethanol precipitates the DNA from the mixed solution since DNA is insoluble in ethanol. Thus, DNA can be seen with the aid of the ethanol. Ethanol is preferred since the pellets are easily dried due to less salt is precipitated (Herzer, 2001). Then, it is centrifuged again for 10 minutes. After centrifuge is done, DNA pellet is observed whether it is visible or not. After that, cold absolute ethanol is discarded by pouring or by using pipette. The DNA pellet is made sure to still in intact at the bottom of the tube.
Next is the DNA washing. During this step, 500µl of cold 70% ethanol is added into the eppendorf tube. The 70% ethanol function is to wash the DNA because the...

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