Assessment of transferability of sorghum (Sorghum bicolor) EST-SSR markers among its wild species and other members of Gramineae family*

assessment of their intraand interspecific/genera transferability. The genomic DNA was extracted using the CTAB method and PCR were performed as described in Srinivas et al., (2008). Number of alleles, allele frequencies, and polymorphic information content (PIC) for each marker was calculated using Powermarker VER. 3.0 (Liu and Muse 2005). Genetic distance matrix was computed using the binary data matrix and the distance matrix was subjected to cluster analysis based on neighbour-joining (Saitou and Nei 1987) algorithm using Darwin 5.0 (Perrier et al. 2003). To confirm transferability of sorghum EST derived markers, BLASTN (Altschul et al., 1990) analysis of SSR-ESTs (http:// www.ncbi.nlm.nih.gov/blast/) with a homology filter of expectation value _ 1.00E-09 was conducted against GenBank data base of rice, maize, sugar cane, pearl millet and oats. Hits with significant Evalue were aligned using ClustalW (Thompson et al. 1994) to observe the conservation of SSR motif. A total of 94 SSR alleles were detected using 14 ESTSSR primer pairs, with an average of 3 alleles/locus in sorghum, and 6.71 alleles in Gramineae as a whole. The number of alleles/primer pair varied from 1 (Stgnhsbm1 and 32) to 6 (Stgnhsbm33) in sorghum, while it was 3 (Stgnhsbm1) to 10 (Stgnhsbm 5, 17 and 33) in Gramineae. The allele size ranged from 95 to 370 bp in sorghum and 95 to 410 bp in Gramineae members. The PIC values varied from zero (Stgnhsbm1 and 32) to 0.74 (Stgnhsbm34) with an average of 0.40 in sorghum, whereas, it ranged between 0.41 (Stgnhsbm1) and 0.85 (Stgnhsbm5, 17 and 34) with a mean 0.69 in Gramineae. All the 14 markers were transferable in S. halepense while 13 markers were PCR amplified in S. usumbarense, parasorghum and S. propinquum (92.8% transferability) indicating robust transferability of S. bicolor EST-SSRs across sorghum species. Of the 14 markers studied, 6 markers (Stgnhsbm 3, 4, 5, 9, 33 and 34) were polymorphic (42.8%) within cultivated sorghums (S. bicolor) and 12 SSR markers are the most popular molecular markers for crop improvement and are frequently used in fingerprinting cultivars, genetic diversity studies, mapping QTL and markerassisted breeding of crop plants. With the availability of large numbers of expressed sequence tags (ESTs) in the public databases, the development of SSR markers has been accelerated to a great extent through in silico approaches with lower cost and effort. Mining of SSR markers from ESTs has been reported in an array of crop plants, including sorghum (Srinivas et al. 2008). Comparative genetic mapping of cereal crops has shown that both gene content and/or order are largely conserved over the evolutionary history of the grasses (Gale and Devos 1998). EST-SSRs, being derived from the conserved expressed component of the genome are expected to show greater transferability between species and genera (Varshney et al. 2005). However, the conserved nature of the EST-SSRs may limit their polymorphism within the species from which they were derived. A relatively high sequence similarity and transferability among members of the Gramineae family have been reported (Kantety et al. 2002, Saha et al. 2004). These markers are useful for assaying functional diversity, comparative mapping and evolutionary studies. The main objective of the present study was to assess the transferability of Sorghum bicolor derived EST-SSR markers to its related wild species, and to other members of Gramineae family. Eighteen genotypes were used in the study (Table 1). A set of 14 random sorghum (S. bicolor) EST-SSRs, viz Stgnhsbm 1, 3, 4, 5, 9, 17, 18, 21, 22, 27, 30, 32, 33 and Stgnhsbm 34 (Srinivas et al. 2008) were used for the