Screening of Indigenous rice by quantitative and molecular characterization for salinity tolerance in Cauvery delta region of Tamilnadu, India.
Description
Genetic diversity using Morpho-Molecular Markers The reserved seeds of all the twenty-eight rice landraces which were collected from the farmers of different areas, were raised in field condition. Genetic diversity among twenty-eight rice genotypes was analyzed using morphological and molecular marker analysis which revealed significant variability for yield and yield component traits. Based on D2 analysis, genotypes were grouped into six clusters, with maximum inter-cluster distance between cluster II and IV. Molecular study: The harvested seeds were saline-treated (4 dSm-1) and the seedling samples taken for DNA extraction and genotyping were done. The number of alleles per locus ranged from 76 in RM 10852 to 252 in the case of RM336. The primer pairs RM 10852, RM 223, RM 10793, RM 10772, RM 493, RM 336, RM 8094, RM 8046, RM 342, RM 3412, RM 1287, RM 413, RM 222, RM 8264, RM 10843 generated amplified products due to amplification of more than one locus. The total alleles identified in the twenty-eight genotypes were classified into two categories: a) shared alleles and b) Unique alleles. A total of 1959 shared bands and 19 unique allele variants were generated in the form of amplified products using 15 primers pair. The overall size of the PCR products amplified using fifteen primer pairs ranged from 70 bps to 350 bps.) Polymorphism Information Content (PIC) Fifteen SSR markers generated 1959 shared alleles and 19 unique alleles with high PIC values indicating polymorphism. The PIC values revealing allele diversity and frequency among the genotypes varied from 0.2633 in the case of RM 3412 to 0.8576 in the case of RM 8046. Considerably, greater magnitude of PIC value was obtained in the primers RM 8046, RM 493, RM 336,RM 10793 and RM 8094. Clustering analysis based on Euclidian distance matrix Clustering analysis based on unweighted pair group method with arithmetic mean (UPGMA) method using DARWIN was separated. Based on this, twenty-eight genotypes under study were grouped into four clusters and twelve sub clusters. The cluster composition is described here under. Cluster1a: Kuzhiyadichan, Anandanoorsanna, Vellaimilagusamba,Anaikomban. Cluster 1b: Soolaikuruvai, Thanga samba Cluster 1c: Kichilisamba Cluster 1d: Kattuyanam Cluster 2a: Perumkoomvazhai, Kalyan samba Cluster 2b: Seempalai Cluster 2c: Neelan samba, Cochin samba Cluster 3a: Mapillai samba, Sanna samba, Sivappukavuni Cluster 3b: Sengalpattu sirumani, Pokkali Cluster 3c: Vellai poovan, Milagu samba, Poovan samba, Sivappusirumani Cluster 4a: Picini,Seeraga samba, Kothamallisamba Cluster 4b: Pachai perumal, Arikavuravi, Poompalai The study demonstrated substantial genetic variability for grain yield per plant, identified marker-trait associations and relationships among traditional landraces of rice. The results provide insights into the diversity that can be exploited for genetic enhancement of saline tolerant rice varieties.
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Methods and Statistical Procedure The Plant Breeding Farm, Annamalai Nagar is situated in 11.39˚ North latitude 79.71˚ East longitude and 12 km away from Bay of Bengal with an altitude of 6 m above sea level. The climate is relatively warm and humid. The average rainfall of Chidambaram received from South West monsoon is 700 mm . Field screening of selected rice land races for their saline tolerance was carried out in Plant breeding farm. The field experiment was laid out in RBD with three replications during samba season 2022-23. Twenty-five days old seedlings were transplanted to the well prepared main field with one seedling per hill. Agronomic and plant protection measures were followed as per recommended for the zone. Five plants were selected from each replication and the observations were recorded on traits viz., days to fifty percent flowering, plant height, flag leaf length, flag leaf breadth, number of tillers per plant, number of productive tillers per plant, panicle length, number of grains per panicle, grain L/B ration, kernel L/B ratio, 1000 grain weight and grain yield per plant . Screening of genotypes for saline tolerance – Laboratory study The harvested seeds of twenty-eight traditional rice landraces were taken and sown in trays separately for normal (distilled water - EC 0.5 dSm¹) and at different levels of salinity (4 dsm¹,8dsm¹ and 12 dsm¹). The experiment was laid out in a Completely Randomized Design (CRD) with three replications. Preparation of saline water for different salinity levels (4 dsm¹, 8dsm¹ and 12 dsm¹). For salinity level 4 dsm¹, saline water was prepared by adding 2.56g of NaCl in one litre of distilled water. For salinity level 8 dsm¹, water solution was prepared by adding 5.12g of NaCl in onelitreof distilled water. For salinity level 12 dsm¹, solution was prepared by adding 7.68g of NaCl in one litre of distilled water. Saline stress is induced on 7th day after sowing by sprinkling saline solutions at different concentration levels (4 dsm¹,8dsm¹ and 12 dsm¹). The observations were recorded for 14 days. Molecular SSR Studies: The leaf samples were collected from the saline- treated (4 dSm-1) seedlings for DNA extraction and genotyping were done as follows. DNA extraction-Quantification of DNA by Nanodrop -Gel electrophoresis-SSR Analysis-PCR - Allele scoring . Analysis of marker data: A closely related diversity measure is the Polymorphism Information Content (PIC). The PIC value for each SSR was estimated by determining the frequency of alleles per locus using the following formula: PIC = 1- Ʃ (Pi)2 Where, Pi is the proportion of samples carrying the ith allele of a particular locus. Statistical analysis : Data obtained from laboratory experiments was statistically analysed using AGRES software. Clustering analysis based on unweighted pair group method with arithmetic mean (UPGMA) method using DARWIN software.