Identification and characterization of traditional rice genotypes for drought tolerance through proteomic approach
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Date
2015
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College of Horticulture, Vellanikkara
Abstract
The study on “Identification and characterization of traditional rice genotypes for drought tolerance through proteomic approach’’ was carried out during 2013-2015. Traditional rice genotypes were collected from RARS, Pattambi and Ambalavayal which was further screened and evaluated for drought tolerance. The identified tolerant and susceptible genotypes was further analysed by 2D gel electrophoresis. Based on the morpho-physiological characters, the plants which showed wilting symptoms were screened according to international standard i.e., IRRI leaf score 7.0. The genotype which showed no wilting symptom even after 90% of the plants were showing wilting was identified as tolerant and the plant which showed early wilting symptom was identified as susceptible genotype. The rice genotypes were transplanted in polybags and water stress was imposed to each genotype at reproductive stage (visual panicle initiation stage). The genotype Chuvanna Modan has taken 25 days to wilt and the genotype Parambuvattan has taken only 10 days for wilting according to IRRI leaf rolling score 7.0. After 90% of plants reached IRRI score 7.0 phenotypic observations were recorded for each genotype. The morphological observations evaluated in each genotype were water mining traits like root length, root volume, root dry weight and root to shoot ratio and the physiological observations evaluated were measurement of Electrical Conductivity (EC), measurement of Stomatal conductance and transpiration rate using Infra red gas analyser (IRGA). The morphological observations recorded for the most tolerant, Chuvanna Modan at IRRI score 7.0 has shown root length, root volume, root dry wt and root to shoot ratio of 92.67 cm, 120 ml, 10.78 g, 0.72 respectively whereas the most susceptible genotype Parambuvattan recorded 93.33 cm, 78.33 ml, 5.35 g, 0.48 respectively. Similarly, the physiological observations like Stomatal Conductance, Transpiration rate and EC for the Chuvanna Modan were 1.11 μ mol of CO2 m-2 s-1, 10.06 m mol of H2O m-2 s-1 and 54.30 d Sm-1 whereas 0.02 μ mol of CO2 m-2 s-1, 0.65 m mol of H2O m-2 s-1 and 108.04 d Sm-1 for Paramabuvattan Leaf samples collected from Chuvanna Modan and Parambuvattan were further subjected for proteome analysis through SDS and 2D gel electrophoresis. The proteins extracted through TCA/Acetone precipitation were analyzed through SDS PAGE and identified a total of 7 differentially expressed proteins. In Chuvanna Modan genotype proteins of size 20 kD, 38 kD and 44 kD were expressed differentially and 34.4 kD protein was down regulated. In Parambuvattan genotype - 20 kD protein was up regulated 23 kD and 33 kD protein was differentially regulated. The differential bands of size 44 kD and 33 kD bands from Chuvanna Modan and Parambuvattan respectively were sequenced by MALDI-TOF/MS (Sandor Proteomics, Hyderabad). 44 kD band was identified as glycosyltransferase At5g03795 isoform X2, 33 kD band as kinesin-like protein NACK1-like isoform X1 and an uncharacterized protein LOC105957228. Many proteins of same molecular weight will be accumulated, in single band of SDS. Hence to highly resolve the protein based on isoelectric point and on molecular weight two dimensional (2D) gel electrophoresis was carried out to differentiate the proteins present in a single band. The total proteins from tolerant and susceptible genotypes were subjected to 2D gel electrophoresis to identify and characterize the up regulated, down regulated and differentially expressed proteins for drought tolerance. Totally four differentially expressed protein spots were identified in Chuvanna Modan. The differential protein spots were sequenced by MALDI-TOF/MS (Sandor Proteomics, Hyderabad). Spot 1 and 2 were identified as ribulose bisphosphate carboxylase small subunit, spot 3 was identified as Protein tyrosine phosphatases and spot 4 was identified as Photosystem II stability/assembly factor HCF136.
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