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Genetic Diversity Analysis in Aus Genotypes Using Ssr Markers(2009-10)

Dr. Md .Shamsher Ali
CSO
Biotechnology Division,Brri-Gazipur

Md.Monirul Islam
SSO
Biotechnology Division,Brri-Gazipur

Dr,Md.Enamul Hoque
CSO and Head (In Charge )
Biotechnology Division,Brri-Gazipur

Dr.Hosneara Hossain
SSO
Biotechnology Division,Brri-Gazipur

S M Hisam Al Rabbi
SO
Biotechnology Division,Brri-Gazipur

Ripon Kumar Roy
SO
Biotechnology Division,Brri-Gazipur

M A Taher
SO
Biotechnology Division,Brri-Gazipur

Abstract/ Executive Summary:

To establish intellectual property right of local Aus genotypes, DNA fingerprinting is necessary. DNA fingerprinting and genetic diversity of local Aus genotypes are studied by SSR markers to protect our local Aus rice genotypes from biopiracy. Diversity analysis of these genotypes by molecular markers is also required to carry out breeding programmes between diverged parents as morphological data are not reliable. In contrast molecular markers can reveal differences among accessions at DNA level and thus give accurate results. Simples sequence repeat (SSR) markers have become the markers of choice for a wide spectrum of genetic population and evolutionary studies to higher degree of polymorphism in a technically efficient and cost-effective way.

Keywords: Genetic Diversity, SSR markers, DNA, AUS genotypes

Location: Boitechnology Division, BRRI-Gazipur

Start Date:

End Date:

Program Area: Variety and Species

Commodity: Rice

Objectives:

To assess extent of genetic variation in traditional and improved varieties.
To establish their relationship for breeding purpose.

Methodology:

Ninety four (six BRRI released and 88 local) Aus genotypes have been screened against 91 SSR markers for this study. DNA was extracted from young leaves of 21 days old plants by mini preparation DNA method (modified CTAB). The concentration of extracted DNA was estimated by Nano Drop Spectrophotometer. Each PCR was carried out in a 10 μL reaction volume containing 1 μL of MgCl2 free 10X PCR buffer, 1.2 μL of 25 mM MgCl2, 0.2 μL of 10 mM dNTPs, 0.2 μL of 5 U/μL Taq DNA polymerase, 0.5 μL of 10 μM forward and reverse primers and 3 μL (10ng) of DNA using a 96 well thermal cycler. The temperature profile use was of 94° C for 5 min (initial denaturation) followed by 35 cycles of 94° C for 1 min (denaturation), 55° C for 1 min (annealing), 72° C for 2 min (extension) with a final extension for 7 min at 72° C. The annealing temperatures were adjusted based on the specific requirements of each primer combination. The PCR products were mixed with gel loading dye and electrophoresed in 8% polyacrylamide gel using vertical polyacrylamide gels for high throughput manual genotyping. Three μL of PCR products were resolved by running gel in 1X TBE buffer for 1.5 - 2.5 hrs depending upon the allele size at around 80-90 volts and 300-500 mA electricity. The gels were stained in 50μl ethidium bromide solution with 500 ml distil water (1 μg/ml ethidium bromide) and documented using UVP RO (Uvipro Platinum, EU) gel documentation unit.

Source of Information: Proceedings of the BRRI Annual Internal Review for 2009-10

Article Link (Online Journal):

Funding Source:

1. Government

Budget:

Total Budget (Tk.) :

Achievement/Findings:

Ninety four (six BRRI released and 88 local) Aus genotypes were screened against 91 SSR markers for DNA fingerprinting. Among the 51 primers showed polymorphism. One hundred and 95 alleles were detected at loci of 51 microsatellite markers across 94 Aus genotypes. Genetic dissimilarity co-efficient was recognized between every two genotypes based on DNA profile. The highest genetic dissimilarity (1.000) was found among the Aus genotypes of Ausa Bogi (Aus2), Aus gram binni (Aus7), Monshi murali (Aus26), Kachilon (2) (Aus44), Panki rat (Aus45), Agaua (Aus46), Chandra moni (Aus69) and Khusni (Aus72). Whereas the lowest genetic dissimilarity was between BRRI dhan42 and BRRI dhan43 (0.1176). Genetic dissimilarity co-efficient revealed that most local Aus genotypes have broad genetic base as they indicated more genetic dissimilarity. On the other hand, BRRI released modern Aus variety showed narrow genetic base as indicated less then low genetic dissimilarity co-efficient. The genetic similarity analysis using UPGMA clustering system generated 11 major genetic clusters at a cut-off similarity coefficient level of 0.37. The highest number of genotypes clustered in group 9 (22) and lowest in group 3 (1) and 6 (1).

Knowledge Management: Report/Proceedings