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DNA Fingerprinting and Genetic Diversity in Aus Rice (oryza Sativa L.) Landraces of Bangladesh

M A Siddique
SSO
Genetic Resources and Seed Division, BRRI, Gazipur

M Khalequzzaman
CSO and Head
Genetic Resources and Seed Division, BRRI, Gazipur

K Fatema
SO
NATP (Phase I)-SPGR project, Genetic Resources and Seed Division, BRRI, Gazipur

M Z Islam
SO
Genetic Resources and Seed Division, BRRI, Gazipur

M H Baktiar
SO
Genetic Resources and Seed Division, BRRI, Gazipur

M M Islam
SSO
Biotechnology Division, BINA, Mymensingh, Bangladesh

Abstract/ Executive Summary:

The allelic diversity and relationships among 48 Aus rice landraces were determined through DNA fingerprinting using microsatellite (SSR) markers. A total of 14 SSR markers for different chromosomes were used to characterize and differentiate the studied rice genotypes. The number of alleles per locus varied from three alleles (RM118) to 18 alleles (RM44) with an average of 9.88. The polymorphic information content (PIC) varied widely among the loci and ranged from 0.3725 (RM107) to 0.9146 (RM519) with an average of 0.7248. The genetic distance-based results found in the UPGMA clustering system revealed six genetic groups with a similarity coefficient of 0.35. Chakila and Shitki saitta had closest distance in the SSR based genetic distance might have same genetic background. Based on genetic coefficient, the diverse landraces Kasalot, Balam, Pankhiraj, Dular, Hashikalmi, Galong, Panbira, Marichbati, Pidi and Surjomoni could be selected as potential parents for varietal improvement programme. The findings of this study should be useful for varietal identification and could be useful for plant breeders in selecting suitable genetically diverse parents for the crossing programme.

Keywords: Aus rice, DNA fingerprinting, Genetic diversity, SSR markers, UPGMA clustering

Location: All over Balgladesh

Start Date:

End Date:

Program Area: Conservation and Biodiversity

Commodity: Plant

Objectives:

To identify the DNA Fingerprinting and Genetic Diversity in Aus Rice (Oryza sativa L.) Landraces of Bangladesh

Methodology:

Forty-eight Aus rice landraces were used in this study. A five gram seed from each of the entry was germinated and then sown in earthen pots for growth and subsequent DNA extraction. Fourteen SSR markers were selected to detect DNA for discriminating the tested Aus rice landraces. Total genomic DNA was extracted from young leaves of three-week-old plants following the simple and modified protocol of Zheng et al., 1995. PCR analysis was performed in 12.5 µl reaction sample containing 5-25 ng of DNA template, 1.25 µl of MgCl₂ free 10X PCR buffer (100 mM Tris-HCl pH 9.0 at 25°C, 500 mM KCl, 0.1% Triton® X-100 and H₂O), 1.5 µl of 25 mM MgCl₂, 0.25 µl of 10mM dNTP, 0.25 µl of 5 U/µl Taq polymerase enzyme, 0.625 µl each of 10 µM forward and reverse primers using a MJ Research single 96-well thermal cycler. The mixture was overlaid with one drop of mineral oil to prevent evaporation. After initial denaturation for five minutes at 94°C, each cycle comprised one min denaturation at 94°C, one min annealing at 55°C, and two min extension at 72°C with a final extension for seven min at 72°C at the end of 35 cycles. The PCR products were mixed with bromophenol blue gel loading dye and were analyzed by electrophoresis on 8% polyacrylamide gel using mini vertical polyacrylamide gels for high throughput manual genotyping (CBS Scientific Co. Inc., CA, USA). 2.5 µl of amplification products were resolved by running gel in 1×TBE buffer for 2-2.5 hrs depending upon the allele size at around 75 volts and 180 mA current. The gels were stained in 0.5 mg/ml ethidium bromide and were documented using UVPRO (Uvipro Platinum, EU) gel documentation unit. Microsatellite or simple sequence repeat (SSR) markers were used for DNA analysis (Temnykh et al., 2001; McCouch et al., 2002). Size for each amplified allele was measured in base pair using Alpha-EaseFC 5.0 software. The summary statistics including the number of alleles per locus, major allele frequency, gene diversity, polymorphism information content (PIC) values were determined using Power Marker version 3.25 (Liu and Muse, 2005). The allele frequency data from Power Marker was used to export in binary format (allele presence=1 and allele absence=0) for analysis with NTSYS- pc version 2.1 (Rohlf, 2002). A similarity matrix was calculated with the Simqual subprogramme using the Dice coefficient, followed by cluster analysis with the SAHN subprogramme using the UPGMA (unweighted pair group method using arithmetic mean) clustering method as implemented in NTSYS-pc.

Source of Information: Bangladesh Rice J. 21 (1) : 59-65, 2017

Article Link (Online Journal):

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

The results obtained from this study provided some useful implications for establishment of Intellectual Property Rights (IPR) issues of Bangladeshi rice landraces. There was a high level of genetic diversity among accessions of Aus rice landraces, suggesting that SSR markers were effective in the detection of polymorphism in this ecosystem. To broaden the genetic base and for the improvement of Aus rice, landraces having the lowest genetic similarities could be selected as parents. Therefore, hybridization should be made between two distant populations. Considering all these criteria and results, the diverse landraces Kasalot, Balam, Pankhiraj, Dular, Hashikalmi, Galong, Panbira, Marichbati, Pidi and Surjomoni could be selected as parents for further breeding programmes. This will bring about greater diversity, which will lead to a high productive index in terms of increase in yield and overall quality.

Knowledge Management: Journal