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Screening of Salt Tolerent Rice Germplasms at the Seedling Stage Using Ssr Markers

R.R. Rajib
Horticulture Division, Bangladesh Institute of Nuclear Agriculture (BINA), Mymensingh-2022

M.M. Islam
Biotechnology Division, Bangladesh Institute of Nuclear Agriculture (BINA), Mymensingh-2022

M.R. Haque
Agricultural Economics Division, Bangladesh Institute of Nuclear Agriculture (BINA), Mymensingh-2022

M.M. Haque
TCP Division, Bangladesh Institute of Nuclear Agriculture (BINA), Mymensingh-2022

Abstract/ Executive Summary:

Twenty rice germplasms were selected for screening salt tolerance at seedling stage in hydrophonic system using nutrient solution and saline water (EC=12 dS/m). Salinized and non-salinized setups were maintained at seedling stage. Germplasms were evaluated individually for salinity tolerance following 1-9 scale using SES of IRRI protocol as well as through SSR markers. In case of phenotyping, out of 20 germplasms 6, 8, 5 and 1 lines were found as salt tolerant, moderately tolerant, susceptible and highly susceptible, respectively using SES. Out of 7 SSR primers three (RM336, RM11757 and RM7075) were utilized to evaluate salt tolerant rice lines. Nine lines were identified as salt tolerant using RM336 marker where RM7075 showed nine and RM11757 showed eight as tolerant. The tested three primers showed 17 lines as heterozygous bands which could be considered as either tolerant or susceptible.

Keywords: Screening, Salt tolerant, Germplasms, SSR markers

Location: Biotechnology Laboratory of Plant Breeding Division, Bangladesh Institute of Nuclear Agriculture (BINA), Mymensingh

Start Date: 00-02-2012

End Date: 00-06-2012

Program Area: Variety and Species

Commodity: Rice

Objectives:

1. To develop salt tolerant rice genotypes using molecular markers, screening of rice germplasms under salinized condition at the seedling stage and evaluation of rice germplasms using microsatellite markers for salt tolerance were done in this study.

Methodology:

Phenotyping the 20 rice germplasms for salinity tolerance: The study was conducted in the glasshouse and Biotechnology Laboratory of Plant Breeding Division, Bangladesh Institute of Nuclear Agriculture (BINA), Mymensingh. Twenty rice germplasm were used for salinity screening at the seedling stage following IRRI standard protocol. All of these germplasms were collected from Bangladesh Institute of Nuclear Agriculture (BINA), Mymensingh. In this system two steps were maintained, one nutrient solution under controlled environment and another was salanized condition maintaining 12 dsm^-1 at seedling stage. Hydroponic system was used at the glasshouse to evaluate for salt tolerance using nutrient solution. Styrofoam seedling floats were required to conduct the experiment. To prepare 1L of nutrient solution, 1.25 ml of each macronutrient and 1.25 ml of micronutrient solutions were mixed with distilled water (dH₂O). The pH of the solution was measured by pH meter. The pH of the solution was monitored regularly and maintained 5.20 to 5.50. Nutrient solution was changed after every eight days. For salinization, crude salt was dissolved with nutrient solution to reach the desired salinity level (12 dS m^-1). The salinity level was measured by EC meter. The modified standard evaluation score (SES) of IRRI was used to assess the visual symptoms of salt toxicity. Initial scoring was started at 15-day after salinization and final scoring was done at 21-day after salinization. Genotyping the 20 rice germplasms for salinity tolerance: Modified CTAB mini prep was used for DNA extraction for 25 day-old seedling (IRRI, 1997). Parental polymorphism survey was done with SSR markers viz., RM28504, RM10701, RM10772, RM585, RM336, RM11757 and RM7075. Out of 7 markers, three polymorphic SSR markers viz., RM11757, RM336 and RM7075 showed polymorphic and clear bands. Each PCR reaction carried out with 15.0 µl reactions containing 1.5 µl 10 X buffer, 0.75 µl dNTPs, 1 µl primer forward, 1 µl primer reverse, 0.5 µl taq polymerase, 8.25 µl ddH₂O and 2.0 µl of each template DNA samples. PCR profile was maintained as initial denaturation at 940C for 5 min, followed by 34 cycles of denaturation at 940C for 1 min, annealing at 550C for 1 min and polymerization at 72⁰C for 2 min; and final extension by 7 min at 72⁰C. Then electrophoresis in 1.5% agarose gel was done after polymorphism in the PCR products and stained in ethidium bromide. Banding patterns were visualized with ultraviolet gel documentation system. The banding patterns of segregating 20 population were scored compared with tolerant and susceptible parents. The segregating population having similar banding pattern to Binadhan-8 were considered as tolerant, similar to Binadhan-7 were considered as salt susceptible and having both banding pattern (Binadhan-8 and Binadhan-7) were considered as heterozygous.

Source of Information: J. Agrofor. Environ. 7 (2): 171-174, 2013; ISSN 1995-6983

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

In case of phenotyping, out of 20 germplasms 6, 8, 5 and 1 lines were found as salt tolerant, moderately tolerant, susceptible and highly susceptible, respectively using SES. Out of 7 SSR primers three (RM336, RM11757 and RM7075) were utilized to evaluate salt tolerant rice lines. Nine lines were identified as salt tolerant using RM336 marker where RM7075 showed nine and RM11757 showed eight as tolerant. The tested three primers showed 17 lines as heterozygous bands which could be considered as either tolerant or susceptible.

Knowledge Management: Journal