Shrama, V.K.Suryaji, Patil Nakul2020-02-042020-02-042019http://krishikosh.egranth.ac.in/handle/1/5810142456An investigation was undertaken to characterize the rice genotypes in relation to grain iron content using the primers designed specifically to target major iron transporter candidate genes and to analyze the significance of iron transporter based target region amplification polymorphism markers in relation to grain iron accumulation in rice genotypes. Using randomized block design with three replications, 28 varieties and advanced breeding lines of rice were evaluated in field and recommended agronomic practices were adopted while raising the crop. Seeds obtained from each entry in each replication after harvesting of the crop were utilized as samples for determination of iron content in unpolished grains. During molecular profile analysis, 18 entries were selected from the two extremes of grain iron distribution range and genomic templates were extracted from two to three weeks old seedlings. Employing six iron transporter candidate genes specific 16 primer pairs, selective amplification of targeted genomic regions was achieved. Molecular profiling was also performed by using a panel of iron transporter based 16 target region amplification polymorphism markers. Exploitable amount of genetic variability was observed in respect of grain iron accumulation amongst 28 varieties and advance breeding lines of rice initially evaluated as experimental materials in the present investigation. While taking into account the range of variation (16.39 to 39.04 ppm) recorded with respect to grain iron concentration, the entries under evaluation were classified into very low, low, moderate, high and very high grain iron containing groups. Grain iron content was observed to be comparatively very high in unpolished grains of Janki, Kanak, Swarna-sub1, RAU-3055 and Vaidehi. Microsatellites, altogether 18 in number with di-nucleotide to penta-nucleotide repeat motifs, were detected within the genomic regions specific to five of the six candidate genes investigated, providing a basis to infer that the sequence length variation present in candidate genes may be a role player in the differential grain iron accumulation in rice varieties. Reproducible amplification in 18 purposefully selected rice varieties and advance breeding lines was successfully achieved by utilization of six candidate genes specific 16 primer pairs during molecular characterization. However, only 12 primer pairs detected targeted genomic regions specific sequence length variation. While three primer pairs generated monomorphic amplified products, one primer pair revealed genetic polymorphism in the form of presence and absence of amplified product. Polymorphic information content of candidate gene specific 12 primers ranged from 0.278 to 0.886 with an average value of 0.540 across the primers. Employing 16 TRAP markers, differential amplification pattern was observed amongst the entries subjected to molecular characterization. Altogether 122 bands including 9 monomorphic bands were scored and categorized into 13 unique alleles and 109 shared alleles. Polymorphic information content of 16 TRAP markers ranged from 0.106 to 0.560 with an average of 0.334 across the markers. Ample molecular level genetic differentiation and divergence was revealed amongst the entries using the candidate genes specific primers and TRAP markers. Hierarchical classification pattern of rice genotypes was almost in complete agreement with principal coordinate analysis based spatial distribution pattern of genetic profiles of rice genotypes. Candidate gene markers based analysis appeared remarkably effective in differentiation of rice genotypes in relation to grain iron accumulation. Hence, these markers can be effectively and efficiently utilized for grain iron accumulation related discrimination of rice genotypes and selection of parental genotypes for genetic improvement in relation to grain iron biofortification. Single marker analysis established the association of two candidate genes specific markers, namely, OsYSL6a and OsNAS2c with grain iron accumulation. These two markers can be effectively used in marker-assisted selection program for grain iron biofortification in rice. Significance of iron transporters based target region amplification polymorphism markers for differentiation and divergence analysis was established in relation to grain iron concentration in rice genotypes. The entries with low and high grain iron content were found to be distributed into different clusters and sub-clusters.ennullThesis