Molecular Characterization of Aerobic Rice Genotypes Using Microsatellite Markers
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Date
2014
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Rajendra Agricultural University, Pusa (Samastipur)
Abstract
study was undertaken to evaluate the nature of microsatellite sites based simple sequence length polymorphism in eighteen aerobic rice entries in order to characterize them on the basis of simple sequence length polymorphism and to determine the nature and extent of genetic differentiation and diversity among them using twenty six microsatellite primer pairs. The statistical methods and parameters used for deriving inference were polymorphism percent, polymorphism information content, discrimination and non-discrimination coefficient, similarity coefficient and numerical taxonomic analysis of divergence.
The amplification was successfully achieved with all the microsatellite primer pairs used in the present study. Altogether 225 allelic variants were detected at 33 microsatellite loci with an average of 6.81 alleles per locus. The number of alleles per locus ranged from five in the case of RM 524 to thirteen in the case of RM 538. A total of 110 shared and 115 unique allelic variants were generated in the form of amplified products by using the 26 primer pairs. The primer pairs RM 538, RM 591, RM 263, RM 335, RM 234, RM 5359, RM 3530, RM 3873, RM461, RM 510, RM 219, RM 332, RM 280, RM 7003 and RM 407 generated considerably higher percentage of unique alleles, clearly reflecting higher polymorphism percentage which was exhibited by these primer pairs in combinations with the entries under evaluation.
The microsatellite locus associated with primer pairs RM 280, RM 3873, RM 114 and RM591 showed null alleles in some of the entries under evaluation. Occurrence of null alleles for a particular repeat locus was noticed reflecting thereby the failure of locus specific microsatellite primer directed generation of amplified
products. While the primer pairs RM 3873, RM 3530, RM 263, RM 234, RM 407, RM 201 and RM 337 generated amplified products due to amplification of more than one microsatellite locus in combination with one to twelve entries under evaluation. Appearance of more than one band in the same entry was noticed revealing the existence of duplicated regions in the genome of rice.
The estimates of polymorphism information content values for the primer pairs varied from 0.607 in the case of RM 407 to 0.913 in the case of RM 591 with an average value of 0.795 per primer pairs. The primer pairs RM538, RM 5359, RM 263, RM 337, RM 591, RM 224, RM 153, RM 3530, RM 332, RM 335and RM 219 appeared to be more informative amongst the primer pairs used for the purpose of molecular characterization of aerobic rice entries under evaluation.
The estimates of discrimination coefficient ranged from 0.647 in the case of primer pair RM 407 to 0.960 in the case of primer pair RM 591. Considerably greater magnitude of discrimination coefficient was obtained in the case of primer pairs RM 591, RM 5359, RM 538, RM 263, RM 335, RM 153, RM 337, RM 224, RM 3530, RM 114, RM 60 and RM 332 in descending order of magnitude. The discrimination coefficient for a primer pair revealed its ability to discriminate the pair-wise combinations of the entries evaluated during the present investigation.
The microsatellite loci with di- nucleotide, tri- nucleotide and complex repeat motifs tended to detect greater number of alleles than the repeat loci with tetra-nucleotide repeat motifs. The results did not indicate a direct relationship between the repeat number involved in the microsatellite locus with di-nucleotide repeat motif and the number of identified alleles. An analysis of allelic diversity data using similarity coefficient based sequential agglomerative hierarchical non-overlapping clustering module indicated that the aerobic rice entries AER-05 and AER-06 were relatively more closely related with the highest similarity coefficient amongst eighteen aerobic rice entries evaluated. The microsatellite marker based analysis revealed unique or entry specific allele which could be useful as DNA fingerprints in the identification and preservation of these aerobic rice entries. The use of twenty six microsatellite markers in the analysis exhibited a remarkably higher level of genetic polymorphism, which allowed unique and unambiguous genotyping of eighteen entries included in the analysis.
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