Isolation of DNA from oral rinse in HPV positive patients

ABSTRACT

To extract human genomic DNA from oral rinse in HPV positive patients. Experimental study. Research Laboratory, Ziauddin Medical University, from February to July 2011. Two hundred and fifty oral rinse samples from human papilloma virus [HPV] positive subjects were collected in 50 ml corning tubes. DNA was extracted from 10 ml of oral rinse by using Lysis buffer [sodium dodecyl sulfate, sodium chloride, sodium citrate and EDTA], isopropanol and 3 M sodium acetate and final washing with ethanol. The DNA was quantified by using Qubit Registered dsDNA BR Assay [Qubit Registered 2.0 invitrogen life technologies USA] and the quality was checked by running an aliquot on 0.6% agarose gel stained with ethidium bromide. PCR were done to amplify beta-globin gene on chromosome 11 by using primers GH20/PCO4 of 260 bp fragment. The mean concentration of all 250 samples DNA was 15.648 +/- 10.50 microg/ml determined by using Qubit Registered 2.0. A single intense band without smearing was seen in almost all cases which confirmed the integrity of DNA. The PCR amplification of human beta-globins primers was successfully done. The oral rinse method was found a simple and highly appropriate means for non-invasive sample collection with easy storage, DNA recovery and subsequent PCR amplification in HPV positive patients