A rapid and inexpensive one-tube genomic DNA extraction method from Agrobacterium tumefaciens.
Many methods have been used to isolate genomic DNA, but some of them are time-consuming and costly, especially when extracting a large number of samples. Here we described an easy protocol using two simple solutions for DNA extraction from A. tumefaciens cells. Compared with the standard protocol, this protocol allows rapid DNA isolation with comparable yield and purity at negligible cost. Following this protocol, we have demonstrated: (1) gDNA extraction was achieved within 15 min; (2) this method was cost-effective, since it only used calcium chloride and lysozyme; SDS, phenol, chloroform and proteinase K were not necessary; (3) the method gave high yield of gDNA (130 ng/loopful culture) compared with standard protocol that was suitable for restriction analysis; (4) the protocol can be carried out in a single test tube and the cells directly from solid media can be used. Thus, this protocol offers an easy, efficient and economical way to extract genomic DNA from A. tumefaciens.
Center for Biotechnology, Pravara Institute of Medical Sciences, Loni, Ahmednagar, Maharashtra, India. suresh.kamble@pmtpims.org.
Suresh P
SP
Kamble
Department of Biochemistry, Dr. Babasaheb Ambedkar Marathwada University, Aurangabad, 411004, Maharashtra, India.
Madhukar M
MM
Fawade
Calcium chloride; Genomic DNA extraction; Lysozyme; Restriction digestion