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DNA Fingerprinting of BRRI Released Varieties (2008-09)

Dr. Md .Shamsher Ali
CSO
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

Md Monirul Islam
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 BRRI released varieties, DNA fingerprinting is necessary. DNA fingerprinting and genetic diversity of BRRI released rice varieties are studied by SSR markers to the protected varieties from biopiracy. Diversity analysis of these varieties by molecular markers is also required to carry out breeding programmes between diverged parents as morphological data are not reliable. Contrastingly molecular markers can reveal differences among accessions at DNA level and thus give accurate results. Single 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. Fifty BRRI released varieties have been screened against 110 SSR markers for DNA fingerprinting. Among them 51 primers showed polymorphism. One hundred and ninety one alleles were detected at loci of 51 microsatellite markers across 50 rice genotypes. Genetic dissimilarity co-efficient was recognized between every two variety based on DNA profile The highest genetic dissimilarity (0.8222) was found between BR5 and BR21. Whereas lowest genetic dissimilarity was between BR22 and BR23 (0.0213). However, most BRRI varieties have narrow genetic base as they showed less genetic dissimilarity co-efficient. The genetic similarity analysis using UPGMA clustering system generated 7 major genetic clusters at a cut-off similarity co-efficient level of 0.49. The highest number of varieties clustered in group 1 and lowest in group 3 and 6.

Keywords: DNA fingerprinting, Markers, Diversity analysis

Location: Biotechnology Division, BRRI-Gazipur

Start Date: 01-01-2008

End Date:

Program Area: Variety and Species

Commodity: Rice

Objectives:

DNA fingerprinting and diversity analysis of BRRI varieties by SSR markers.

Methodology:

Fifty BRRI released varieties have been screened against 110 SSR markers for this study. DNA was extracted from young leaves of 21 days old plants by mini DNA preparation 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 2 μL (10ng) of DNA using a 96 well thermal cycler. The temperature profile use for PCR amplification comprised 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 at the end of 35 cycles. 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. Four μL of amplification products were resolved by running gel in 1X TBE buffer for 1.5 - 2.5 hrs depending upon the allele size at around 75 volts and 180 mA electricity. The gels were stained in 1 μg/ml ethidium bromide solution and were documented using UVPRO (Uvipro Platinum, EU) gel documentation unit.

Source of Information: Proceedings of the BRRI Annual Internal Review for 2008-09

Article Link (Online Journal):

Funding Source:

1. Government

Budget:

Total Budget (Tk.) :

Achievement/Findings:

Most BRRI varieties have narrow genetic base as they showed less genetic dissimilarity co-efficient. The genetic similarity analysis using UPGMA clustering system generated 7 major genetic clusters at a cut-off similarity co-efficient level of 0.49. The highest number of varieties clustered in group 1 and lowest in group 3 and 6.

Knowledge Management: Report/Proceedings