EFFECT OF PROTEINASE K AND ELUTION BUFFER ON DNA QUALITY OF PRIMARY SEX ORGANS OF KUNDHI BUFFALO
Keywords:buffalo, DNA, porteinase K, testes
A study was conducted to optimize a rapid and cheaper mammalian DNA extraction protocol and to evaluate the effect of extracted DNA stored on various temperatures. Hundred fresh tissues of primary sex organs of Kundhi buffalo were collected from local slaughter house. The DNA was extracted using commercial kit (protocol-1), its modified methods (protocol 2-4) and phenol chloroform method, the fresh and stored DNA was measured through spectrophotometer. The concentration of DNA extracted from ovary by using commercial kit and its modified methods of protocol 2, 3, 4 and phenol chloroform method was 40.9, 42.2, 88.7, 226.2 and 232.7 ng/µl, respectively and with similar protocols in testes the concentration of extracted DNA was 41, 34.6, 64, 169.7 and 233.3 ng/µl, respectively. Significantly (P?0.001) higher concentration of DNA was obtained through modified methods of commercial kit and in phenol chloroform method than normal recommended protocol kit. The average purity (absorbance ratio A260/A280) from both primary sex organs was higher than 1.5 and ranged between 1.6-1.8 in ovary, whereas in testes it ranged between 1.6-2.0, in both of the organs. The higher absorbance ratio was measured through phenol chloroform method of DNA extraction. In conclusion the phenol chloroform and commercial kit methods yielded a good quality DNA and may be used for further experiments, like PCR related down streaming techniques. Phenol chloroform method is laborious but cost effective and may be used as an alternative of commercial kit for DNA extraction from primary sex organs of Kundhi buffalo.
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