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Genetic Diversity in Aus Rice Genotypes Using Ilp Markers

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

M Khalequzzaman
Genetic Resources and Seed Division, BRRI, Gazipur, Bangladesh

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

E S M H Rashid
Genetic Resources and Seed Division, BRRI, Gazipur, Bangladesh

M H K Baktiar
Genetic Resources and Seed Division, BRRI, Gazipur, Bangladesh

M A Z Chowdhury
Member Director (Crops), BARC, Dhaka, Bangladesh

Abstract/ Executive Summary:

Assessment of genetic diversity is essential for germplasm characterization, utilization and conservation. Genetic diversity of 31 Aus rice genotypes of Bangladesh was assessed using 11 ILP (intron length polymorphism) markers. A total of 28 alleles were detected and the number of alleles per locus varied from 2 (RI01779, RI05751, RI05304, RI03205, RI00299, RI05407) to 4 (RI05559). The PIC values ranged from 0.06 (RI05407) to 0.57 (RI05559) with an average of 0.33. PIC value revealed that RI05559 was the best ILP markers for the studied 31 Aus rice genotypes. The dendrogram from unweighted pair-group method with arithmetic average clustering classified the genotypes into five groups at a coefficient of 0.57. Two dimensional graphical views of Principal Coordinate Analysis (PCoA) revealed that the genotypes Kuchmuch, Kalo dhan, Aus dhan, Sadey aus, Chaina and Dighi bawalia were found far away from the centroid of the cluster and can be seslected as parents for further breeding programmes. Parangi and V3, Adubali and H1-2, Begunbichi and Hashikalmi had closest distance (0.000) in the distance matrix might have same genetic background. This information will be useful for the selection of genetically diversed parents and assist in trait development using genotypes in rice breeding programmes in future. The results provided some useful implications for establishment of sovereignty of Bangladeshi rice gene pool. It was also suggested that ILP markers could be very useful for the genetic study and breeding in rice.

Keywords: Aus rice, Genetic diversity, Cluster analysis, Allele, Polymorphism

Location: Genetic Resources and Seed Division of BRRI, Gazipur, Bangladesh

Start Date: 00-00-2016

End Date: 00-00-2016

Program Area: Conservation and Biodiversity

Commodity: Plant

Objectives:

To determine the Genetic Diversity in Aus Rice Genotypes using ILP Markers

Methodology:

Thirty-one Aus rice genotypes were studied in the Molecular Laboratory of Genetic Resources and Seed Division of BRRI during 2016 for diversity analysis. Five gram seeds from each of the entry were first germinated and then sown in the earthen pots. Eleven ILP markers (synthesized by Beijing Sunbiotech Co., Ltd., China) were used for diversity analysis. DNA was extracted from young leaves of 14-day-old plants following the Miniscale method (Zheng et al., 1995). The total PCR reaction volume was 10 μL, composed of 3.0 μL genomic DNA, 1.0 μL of 10 × PCR buffer (MgCl₂ free), 1.35 μL of 25 mmol/L MgCl₂, 0.2 μL of 10 mmol/L dNTPs, 0.5 μL of 10 μmol/L forward and reverse primers, 0.02 μL of 5 U/μL Taq DNA polymerase and 3.43 μL sterile deionized water. The temperature profile was an initial denaturation step for 5 min at 94°C, followed by 35 cycles of denaturation (94°C) for 45 s, annealing (55°C or 61°C) for 45s and primer elongation (72°C) for 1.3 min and then a final extension at 72°C for 7 min. The 10 μL of PCR product with 2 μL of a loading dye was analyzed using 8% polyacrylamide gel electrophoresis in 1 × TBE buffer at 75 V for about 2.0–2.5 h depending upon the allele size. The gels were stained with ethidium bromide solution (0.5 mg/mL) for 25 min and exposed to UV light using the molecular imager gel documentation unit (XR System, Uvitec Cambridge, France) for visualization. The size of the band for each marker was scored by Alpha-EaseFC 4.0 software. The summary statistics, including the number of alleles, major allele size and frequency, gene diversity and polymorphism information content (PIC) value, were determined using PowerMarker version 3.25 (Liu and Muse, 2005). For the unrooted phylogenic tree, genetic distance was calculated using the “C.S. Chord 1967” distance (Cavalli-Sfoza and Edwards, 1967) followed by phylogeny reconstruction using neighbour-joining as implemented in PowerMarker with tree viewed using Tree view (Page, 1996). The allele frequency data from PowerMarker was used to export the data in binary format (allele presence=“1” and allele absence = “0”) for analysis with NTSYS- pc version 2.2 (Rohlf, 2002). A similarity matrix was also calculated with the Simqual subprogramme using the Dice coefficient followed by cluster analysis with the SAHN subprogramme using the UPGMA (Un- weighted pair group method using arithmetic average) clustering method as implemented in NTSYS-pc. The similarity matrix was also used for principal coordinate analysis (PCoA) with the DCenter, Eigen, Output, and MXPlot subprogrammes in NTSYS-pc.

Source of Information: Bangladesh Rice J. 20 (2) : 13-19, 2016

Article Link (Online Journal):

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. There was a high level of genetic diversity among Aus rice genotypes in this study, suggesting that ILP markers were very effective in the detection of polymorphism in this ecosystem. To broaden the genetic base and improvement of Aus rice, genotypes having the lowest genetic similarities could be selected as parents. Therefore, hybridization should be made between two distant populations. Based on distance matrix, the genotypes Kuchmuch, Kalodhan, Kalohizli, Sandamoni, Kalo aus, Boula, Minikit, Begunbichi, Hanumanjata and Dighibawalia can be selected as parents for further breeding programmes. Parangi and V3, Adubali and H-12, Begunbichi and Hashikalmi had closest distance (0.000) in the distance matrix might have same genetic background. The findings of this study should be useful for varietal identification and could help in providing guidelines for assisting rice breeders in selecting suitable genetically diverse parents for the crossing programme.

Knowledge Management: Journal