Evaluation of Easy and Economic Protocols for High Quality Genomic DNA Extraction and Sex Determination in Papaya Using SCAR Markers

Abstract

Rapid and precise identification of three sex types (male, hermaphrodite and female flowers) is the key in achieving good returns in papaya (Carica papaya L.) cultivation. This study presents a simple, reliable, fast and cost-efficient DNA isolation protocol and a rapid and precise method of sex determination in papaya. Three DNA isolation protocols viz., CTAB method, Modified Dellaporta method and BioBasic DNA isolation kit were comparatively evaluated for their simplicity, economic, DNA quantity and purity. Among them, CTAB method was regarded as relatively economic, with a cost of Rs. 11.80943 per sample with higher amount of DNA (4667.4 µg/ µl). However, it was noticed that DNA isolation with the BioBasic kit was fastest method, which took only 6 hours 7 minutes. On the other hand, relatively better DNA purity was noticed with the Modified Dellaporta method  (average ratio of absorbance at 260 nm and 280 nm range between 1.8-2.0) and yielded clear, intact and distinct DNA  bands. Hence, Modified Dellaporta method is suggested as an ideal DNA isolation protocol for papaya. Molecular markers (such as SCAR derived from RAPD T12, W11, NAPF-2 and PKBT-4) were employed in this study to identify the sex type and they have successfully distinguished between male and hermaphrodite plants in both dioecious and gynodioecious varieties; especially, NAPF-2 has successfully distinguished male in dioecious varieties. Evaluation of different kinds of molecular markers has shown that the SCAR marker T12 can be used as reliable marker for rapid and precise identification of papaya sex type.