Safflower (Carthumus tinctorus L.) is a flowering annual plant belonging to Asteraceae family. It is one of the oldest crops known to human beings in the world grown for centuries in India.
Safflower cultivation in India is constrained by the susceptibility of cultivar germplasm to an array of biotic and abiotic stresses, such as wilt, root rot, Alternaria leaf spot, aphids, salinity and alkalinity. Sources of resistance to these stresses are rather limited in the cultivar germplasm. Hence there is need to introgress the desirable genes from wild species. Before undertaking introgressive breeding there is a need to find out the degree of genetic relatedness of cultivated safflower to wild Carthamus species. In view of this the present investigation is aimed at
1. Collection, establishment and characterization of Carthamus species
2. Karyomorphological characterization of Carthumus species
3. Assessment of nuclear genomic diversity using random primers like Random Amplified Polymorphic DNA (RAPD) and Inter Simple Sequence Repeats (ISSR).
One cultivated safflower variety; eleven wild species with three accessions were used in this study. These species were characterized by both morphological as well as karyomorphological (cytological and ploidy level by ploid analysis) means. The species under study had basic chromosome numbers of 10, 12, 22 and 32. The 2C-DNA content of the species varied between 45 (C. boissieri) and 297 (C. creticus)
The genetic variation in the nuclear genome of these wild Carthamus species was assessed through 2 types of PCR based genetic markers. Viz.,RAPD and ISSR. Following initial screening of 120 random decamer primers for amplification and polymorphism, 27 primers which gave good amplification and informative fingerprints were selected for diversity analysis. Estimates of genetic similarity based on Dice coefficient ranged from 0.32 (32%) to 0.93 (93%) indicating greater genetic variability among the safflower species based on RAPD markers. Among the species studied C. creticus was distantly placed to cultivated safflower, C. tinctorius. Despite the high degree of polymorphism generated the RAPD markers grouped accessions of the same species in separate clusters. Hence RAPD markers should be used as a caution for classification.
Based on diversity analysis data generated through RAPD polymorphism, two species viz., C. tinctorius and C. creticus were selected for an initial screening of 100 heterologus ISSR primers. Of these 31 showed amplification in either one or both the species. From these 11 primers which showed good amplification and polymorphism were selected for further analysis. Estimates of genetic similarity based on Dice coefficient ranged from 0.26 (26%) to 0.94 (94%) indicating greater genetic variability among the safflower species based on ISSR markers and was on par with the polymorphism generated by RAPD markers. Unlike RAPD polymorphism, with ISSR primers the cultivated species and its closest relatives formed one cluster whereas distant wild species formed separate clusters. The high diversity detected by ISSR marker is consistent with the known characteristics that they are effective multilocus markers for applications such as diversity analysis, fingerprinting and genome mapping. The results based on those two marker systems suggests that RAPD markers could be used to assess the intra varietal variation while ISSR markers could be an useful tool in phylogenetic analysis.
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