A Protocol for Obtaining PCR-Quality DNA from Formalin Preserved Cysts of Potato Cyst Nematode Globodera rostochiensis

Preservation of biological samples in formalin causes formation of DNA-protein cross linking and DNA fragmentation. Obtaining PCR-quality genomic DNA from formalin fixed nematode samples is a tricky process. In this study, we demonstrate

obtaining of PCR-quality DNA from formalin preserved cyst of potato cyst nematode Globodera rostochiensis by increased and extended

washing, followed by lysis by modified protocol. The quality of extracted DNA was confirmed by PCR amplification using G. rostochiensis species-specific primers, and a sharp 318bp sized band was resolved on 1.2% agarose gel. Using the modified protocol, we succeeded in getting a positive PCR amplification in 54–63% formalin fixed nematode specimen. The PCR amplification sensitivity for formalin fixed sample was found up to 10⁻³ dilution from crude DNA extracted from a single cyst. To the best of our knowledge, this is the first protocol to extract PCR-quality DNA from formalin fixed sample of potato cyst nematode G. rostochiensis. This protocol may also be used on other formalin fixed nematode samples.