Submitted to: BMC Research Notes
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/18/2008
Publication Date: 6/18/2008
Citation: Barkley, N.L., Wang, M.L., Gillaspie Jr, A.G., Dean, R.E., Pederson, G.A., Jenkins, T.M. 2008. Discovering and Verifying DNA Polymorphism in a Mung Bean [V. radiata (L). R. Wilczek] Collection by Ecotilling and Sequencing. Biomed Central (BMC) Notes 1:28. Interpretive Summary: Vigna radiata, which is classified in the family Fabaceae, is an important economic crop and a dietary staple in many developing countries. The species radiata is further subdivided into varieties such as radiata and sublobata (both of which are part of the USDA germplasm collection), of which sublobata is currently acknowledged as the putative progenitor of radiata. Previous molecular genetic studies as well as collected morphological descriptors such as flowers, and pod characters of V. radiata have demonstrated limited genetic diversity in the USDA germplasm collection. Ecotilling, a relatively new molecular procedure was employed to rapidly mine for DNA variation. This technique allows numerous samples to be processed efficiently and the variation between samples to be quickly identified compared to other molecular techniques. The Ecotilling method demonstrated to be highly effective at uncovering genetic variation between radiata and sublobata. It also was beneficial in identifying genetic variation among radiata accessions. Uncovering this genetic variation can provide information on the function of a gene or be beneficial by designing markers linked to a gene of interest which can be used in breeding for specific traits.
Technical Abstract: Ecotilling was used to identify various SNPs and small insertion/deletions (INDELS) in a collection of mung bean Vigna radiata, which was previously shown limited diversity. Ten primer sets were used to examine intervarietal and intravarietal DNA polymorphisms among various mung bean accessions and its wild relative. Initially, Vigna radiata var. sublobata DNA was used as a reference and compared to all members of the population V. radiata for DNA polymorphisms. A total of 157 DNA polymorphisms were observed in the population when using V. radiata var. sublobata as the reference. The banding patterns varied from simple to complex as the number of DNA polymorphisms between two pooled samples increased. Numerous SNPs and INDELS ranging from 4-24 and 1-6, respectively, were detected in all PCR fragments when comparing V. radiata var. sublobata to V. radiata var. radiata. Additionally, accessions in the population were pooled together to identify SNPs and INDELS among V. radiata var. radiata. When accessions of V. radiata were mixed together and digested with CEL I, very few SNPs and INDELS were detected, suggesting that radiata has limited genetic diversity. Morphology data taken from these accessions of V. radiata also demonstrated limited diversity in the flowers and pod descriptors. Ecotilling is a beneficial technique for efficient identification of DNA polymorphorism in populations with high genetic identity and to mine for SNPs in plant germplasm. The Ecotilling technique, beneficial applications, and limitations are discussed.