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Molecular Characterization and Genetic Diversity in Geographical Indication (gi) Rice (oryza Sativa L.) Cultivars of Bangladesh

M. A. Siddique
GRSD, BRRI, Gazipur 1701, Bangladesh

M. Khalequzzaman
GRSD, BRRI, Gazipur 1701, Bangladesh

M. M. Islam
IAPP-BRRI, Barisa¨l, Bangladesh

Kaniz Fatema
NATP-SPGR Project, GRSD, BRRI, Gazipur 1701, Bangladesh

M. A. Latif
Plant Pathology Division, BRRI, Gazipur 1701, Bangladesh

Abstract/ Executive Summary:

Assessment of genetic diversity and molecular characterization among elite rice cultivars of Bangladesh is important for varietal identification. Genetic diversity of 20 geographical indications (GI) rice landraces of Bangladesh was analyzed for 30 loci using simple sequence repeat markers to characterize the varieties and also to establish the sovereignty of the Bangladeshi rice gene pool. The number of alleles per locus ranged from five (RM275) to 15 (RM180), with an average of 9.7. The results revealed unique alleles that could be used for the identification and molecular characterization of five landraces. The polymorphism information content (PIC) values which ranged from 0.59 (RM275) to 0.90 (RM180), with an average of 0.815, revealed many variations among the studied cultivars. The frequency of the most common allele at each locus ranged from 15 (RM85 and RM180) to 55 % (RM275 and RM277). RM180 was the best marker for identification and diversity estimation of GI rice genotypes as revealed by PIC values. A dendrogram revealed six clusters with a similarity coefficient of 0.14. Two- and three-dimensional graphical views of the principal coordinate analysis showed the spatial distribution of the genotypes. The cultivars Galon, Radhunipagal, Biruin, Joina, Chamara, Horkoch, Sadamota, Topa, Kalijira, Binni, Balam, and Kataribhogh were found far away from and distributed around, the centroid of the cluster. The findings of this study are useful for varietal identification, thus assisting plant breeders in selecting suitable genetically diverse parents for crossing programs.

Keywords: Genetic diversity, Geographical indication (GI), Landrace, Microsatellite markers, Rice (Oryza sativa L.)

Location: GRSD, BRRI, Gazipur 1701, Bangladesh

Start Date:

End Date:

Program Area: Variety and Species

Commodity: Rice

Objectives:

The objectives of this research were to (1) assess the genetic variation and diversity of 20 GI rice genotypes and (2) determine the genetic relationship among these genotypes for breeding purposes.

Methodology:

Plant materials Twenty GI rice landraces of Bangladesh were studied. A 5-g seed from each of the entries was germinated and then sown in earthenware pots for growth and subsequent DNA extraction. Genotyping protocol Total genomic DNA was extracted from young leaves of 3-week-old plants following the simple and modified protocol of Zheng et al. (1995). PCR analysis was performed in 12.5 ll reaction sample containing 5–25 ng of DNA template, 1.25 ll of MgCl₂-free 109 PCR buffer (100 mM Tris–HCl, pH 9.0 at 25 C, 500 mM KCl, 0.1 % Triton X-100, and H₂O), 1.5 ll of 25 mM MgCl2, 0.25 ll of 10 mM dNTP, 0.25 ll of 5 U/ll Taq polymerase enzyme, and 0.625 ll each of 10 lM forward and reverse primers using an MJ Research single 96-well thermal cycler. The mixture was overlaid with one drop of mineral oil to prevent evaporation. After initial denaturation for 5 min at 94 C, each cycle comprised 1-min denaturation at 94 C, 1-min annealing at 55 C, and 2-min extension at 72 C with a final extension for 7 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 ll of amplification products were resolved by running gel in 19 TBE buffer for 2–2.5 h depending upon the allele size at around 75 V and 180 mA. The gels were stained in 0.5 mg/ml ethidium bromide and were documented using UVPRO (Uvipro Platinum, EU) gel documentation unit. Data analysis Molecular weight for each amplified allele was measured in base pair using Alpha-Ease 5.0 software. The summary statistics including the number of alleles per locus, major allele frequency, gene diversity (the total number of genetic characteristics in the genetic makeup of a species), and polymorphism information content (PIC) values were analyzed using PowerMarker-3.25 (Liu and Muse 2005). For the unrooted phylogenetic tree, genetic distance was calculated using the ‘‘C.S. Chord 1967’’ distance measure (Cavalli-Sforza and Edwards 1967) followed by phylogeny reconstruction using neighbor-joining as implemented in PowerMarker using TreeView (Page 1996). The allele frequency data were exported in binary format (allele presence = ‘‘1’’ and allele absence = ‘‘0’’) for analysis with NTSYS-pc version 2.2 (Rohlf 2002). A similarity matrix was calculated with the SimQual subprogram using the Dice coefficient, followed by cluster analysis with the SAHN subprogram using the UPGMA (unweighted pair group method using the arithmetic mean) clustering method as implemented in NTSYS-pc. The similarity matrix was also used for principal coordinate analysis (PCA) with the center, Eigen, Output, and MXPlot subprograms in NTSYS-pc.

Source of Information: Braz. J. Bot

Article Link (Online Journal): DOI 10.1007/s40415-016-0271-1

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

The results obtained from this study on molecular characterization provided some useful implications for establishment of sovereignty of Bangladeshi rice gene pool. Considering results from marker-assisted diversity analysis, accessions that are far apart based on their genetic coefficient (like Balam and Kalijira; Biruin and Joina; Kalijira and Bashful; Radhunipagal and Biruin) could be selected as parents for further breeding programs. 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