Abstract
Extraction of high-quality DNA from Gossypium (Cotton) species is notoriously difficult due to high contents of polysaccharides, quinones and polyphenols other secondary metabolites. Here, we describe a simple, rapid and modified procedure for high-quality DNA extraction from cotton, which is amenable for downstream analyses. In contrast to other CTAB methods, the described procedure is rapid, omits the use of liquid nitrogen, phenol, CsCl gradient ultracentrifugation, uses inexpensive and less hazardous reagents, and requires only ordinary laboratory equipment. The procedure employed the high concentration of NaCl and use of PVP-10 to rid the problems associated with polysaccharides and polyphenols, respectively. The average yield was approximately 10–15 μg of good quality DNA from 100 mg of tissue weight, which is adequate for projects, like genetic mapping or marker-assisted plant breeding. This protocol can be performed in as little as 3 h and may be adapted to high-throughput DNA isolation.•Buffers A and B were redesigned from Paterson et al. (1993) and Porebski et al. (1997), respectively.•Ribonuclease A was added before chloroform extraction.•A simple, rapid and inexpensive DNA extraction method is described.
Highlights
Extraction of high-quality DNA in sufficient quantity is important for studying the molecular genetics of cotton
Over the years to overcome these problems, numerous modifications have been introduced into the original Cetyltrimethylammonium bromide (CTAB) method [13] to reduce the cost and time of routine DNA isolation [14]; none of the modifications have been found to be universally applicable for every plant species due to their chemotypic heterogeneity
The described procedure was modified from Paterson et al [9] to reduce time, cost, and resolve the problems associated with high endogenous levels of secondary metabolites, especially polysaccharides and polyphenols
Summary
Qasim Alia,*, Ibrahim Bala Salisua,b, Ali Razac, Ahmad Ali Shahida, Abdul Qayyum Raoa, Tayyab Husnaina a Centre of Excellence in Molecular Biology, University of the Punjab, Lahore-54000, Pakistan b Department of Animal Science, Faculty of Agriculture, Federal University Dutse, P.M.B. 7156, Jigawa State, Nigeria c Centre of Agricultural Biochemistry and Biotechnology, University of Agriculture, Faisalabad-38040, Pakistan
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