Characterization of Groundnut (Arachis hypogaea L.) varieties through DNA Fingerprinting, Isozymes and Protein Profiling.
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Date
2011-05
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jau,junagadh
Abstract
Groundnut (Arachis hypogaea L., 2n=40, Family: Fabaceae) is an important crop for the production of oil. Three DNA based molecular marker techniquesviz.Random Amplified Polymorphism DNA (RAPD), Inter Simple Sequence Repeat (ISSR) and Simple Sequence Repeat (SSR), were used to study the genetic diversity in groundnut varieties.Peroxidase, Esterase, Polyphenol oxidase and Superoxide dismutase Isozymepatterns and protein profiling were used for biochemical characterization of groundnut varieties. Out of the 30RAPD,10 ISSR and 10 SSR primers screened, a total of 39 polymorphic primers (22 RAPD, 9 ISSR and 8 SSR) were used for further study. Amplification of genomic DNA of 12varieties, using RAPD analysis, yielded 96 polymorphic fragments with 100 % polymorphism. Number of amplified fragments with RAPD primers ranged from 3 to 10 and varied in size from 144 to 3315bp. A dendrogram based on UPGMA analysis grouped the 12 groundnut varieties into two main clusters named A and B, with Jaccard’s similarity coefficient ranging from 0.274 to 0.726. The 9 ISSR primers produced 58 bands across 12groundnut varieties, which were polymorphic. The size of amplified bands varied from 153 to 915bp. A dendrogram based on UPGMA analysis of 12 groundnut varieties generated by pooled ISSR molecular data formed two main clusters, named A and B and Jaccard’s similarity coefficient ranged from 0.414 to0.789. The 8 SSR primers produced 15 bands across 12groundnut varieties, which were polymorphic. A dendrogram based on UPGMA analysis of 12 groundnut varieties generated by pooled SSR molecular data formed two main clusters, named A and B and Jaccard’s similarity coefficient ranged from 0.142 to 0.717.
Isozymes patterns werecarried out to assess the biochemical character in groundnut varieties. Four enzyme systems viz., esterase, peroxidase, polyphenol oxidase and superoxide dismutase were studied at different days after germination. Diverse banding pattern was obtained at 6 days after germination (DAG) and9 DAG. Total 6 bands of esterase isozyme were observed at 6 and 9 DAG. Relative mobility of 6 DAG varied between 0.146-0.283 while at 9 DAG it varied from 0.580-0.648.Polymorphism, observed was 100%, and 66.67 % at 6 and 9 DAG, respectively. Total 14 bands of peroxidase isozyme were observed at 6 and 9 DAG. Relative mobility at6 DAG varied between 0.196-0.570; at9 DAG it varied from 0.084-0.508. Polymorphism observed was62.50 %and33.33 % at 6 and 9 DAG, respectively. Total 8 bands of polyphenol oxidaseisozymes were observed at 6 and 9 DAG. Relative mobility of 6 DAG varied between 0.209-0.497 while at 9DAGvaried from 0.261-0.684. Polymorphism observed was 66.67 %and 60 % at 6 and 9 DAG, respectively. Total 8 bands of superoxide dismutaseisozymes were observed at 6 and 9 DAG. Relative mobility of 6 DAG varied between 0.451-0.570 while at 9DAG varied from 0.490-0.735. Polymorphism observed was 0 %and 50 % at 6 and 9 DAG, respectively.
Total of 14 bands were generated by protein profiling using polyacrylamide gel electrophoresis inwhich 4 were polymorphic and showing 66.67 % polymorphism. Overall PIC value for all groundnut varieties reveled by protein profiling was 0.821.The dendrogram constructed based on genetic distance revealed that twelve groundnut varietiesfell into two main clusters,named A and B.
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biotechnology