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Use of Bulk Segregant Analysis for Varietal Identification and Genetic Diversity Estimation in Pakistani Basmati and Non-Basmati Rice Cultivars through Molecular Markers

Arif M, Waheed R, Muqaddasi QH, Ahmed Z, Khalid M, Naveed SA, Shahzad M and Xu J

Identification of alleles for a particular variety through Bulk Segregant Analysis (BSA) is a valuable technique that probes differences specifically in distinguishing traits. Our study was an attempt to develop variety specific marker which differentiate aromatic rice cultivars from non-aromatic. Bulk Segregant Analysis (BSA) approach with RAPD and ISSR assay was undertaken to trace the variety specific alleles among 2 group of popular Pakistani Basmati and non-Basmati rice genotypes to check adulteration by rice exporters. Out of 160 RAPDs and 30 ISSR used in BSA, 29 RAPD and 18 ISSR revealed consistent polymorphisms and were used to assort the advanced disparities amongst 10 varieties. Overall 262 of 359 random amplified alleles and 116 of 151 inter simple sequence alleles were found polymorphic. The number of alleles generated from RAPD and ISSR marker ranged from 5 to 19 and 3-18 respectively. Pair-wise genetic similarity among the varieties with jaccard coefficients was 0.85 for RAPDs and 0.79 for ISSRs. UPGMA dendrogram based on cluster analysis of individual and combine genetic similarity co-efficient resolved 10 rice cultivars into 2 major groups, Basmati and Non-basmati. Moreover, a total of 3 variety specific alleles of 650, 800, 1075 bp were amplified by random primers OPN-05 and OPE-09. While ISSR generated five (UBC-814, UBC-811, UBC-808, UBC-808a and UBC-82) rare alleles for specific genotypes with band sizes of 1500, 1400, 250, 950 and 720 bp respectively. Direct sequencing of rare allelic amplified bands in Sequence Characterized Amplified Regions (SCARs) and sequence tagged sites (STS) might behave useful for recreating phylogenic trees with aim of preserving the integrity of Basmati rice.