Identification of a robust protocol for isolating PCR amenable DNA from caryopses to employ DNA fingerprinting to claim the plant breeders’ rights and varietal ownership of the exporting rice bulks

Abstract

Rice Research and Development Institute (RRDI) at Bathalagoda and its' substations are solely conducting rice breeding in Sri Lanka. Recently, RRDI has identified five exportable rice varieties/lines. As rice export in Sri Lanka is undertaken by the private sector, the RRDI does not receive any foreign revenues or credit for breeding rice genotypes. Therefore, RRDI currently is in need of a reliable protocol to claim the Plant Breeders' Rights (PBR) of these exportable genotypes. The first step of establishing such a protocol based on DNA fingerprinting is depending on the ability to extract good quality DNA from rice caryopses. Therefore, the present study compared the suitability of commonly available DNA extraction methods to purify DNA from rice caryopsis. Three rice genotypes; Bg 250 and At 353 (varieties for local consumption) and Bw-Bs-1-2-31 (with export potential) were selected. Three parboiled rice samples each from long-and short-grain groups were also selected. The DNA extraction, quantification and purity assessment were carried out using four methods viz., CTAB, modified CTAB, Promega kit and Qiagen kit. The extracted DNA was subjected to simplex and multiplex PCR using four rice specific DNA markers. Although positive PCR bands were obtained for all four methods, the modified CTAB method yielded the significantly highest DNA yield (6196.67µg/ml) compared to other methods. Thus, RRDI can employ modified CTAB method in the protocol to claim its PBR and rice varietal ownerships.