PCR-compatible genomic DNA isolation from different tissues of rice (Oryza sativa) for SSR fingerprinting

Abstract

Background: In the genomic era, polymerase chain reaction (PCR) based DNA marker analysis is widely used for several crop plants, especially in rice (Oryza sativa), for several improvemental aspects. Such study requires a fast, inexpensive, and suitable DNA isolation protocol having several overall advantages. The aim of this work was to standardize a DNA isolation protocol for rice which should be simple, cost-effective, high throughput, PCR compatible, and needs a small amount of plant tissues without using liquid nitrogen. Materials and Methods: To fulfill such a desired goal, genomic DNA was isolated from different tissues (seedling, leaf, root, grain, kernel, straw, and embryogenic callus) of the rice plant following a modified protocol. The isolated DNA was subjected to PCR amplification with a reported trait linked rice-microsatellite (RM) marker. Results: The quality and quantity of the isolated genomic DNA from this modified protocol proved to be comparable with the other standardized protocols. The microsatellite based DNA fingerprint shows reproducible bands from different isolated DNA tissue sources. Conclusions: This mini prep cost-effective protocol was standardized with few milligrams of fresh and dried tissues, it does not require liquid nitrogen, can handle large number of samples in a working day per worker, and be efficiently applied to rice. The protocol has now also been applied in other plants like wheat and mungbean yielding about 0.55 μg of high molecular weight DNA from 100 mg of plant material with negligible RNA contamination.