Construction of a Microsatellite DNA Library and Isolation of Microsatellite Loci in Tibetan Chicken

A microsatellite-enriched genomic library for the Tibetan chicken was constructed by the method of enrichment by magnetic beads,and some Tibetan chicken microsatellite sequences were isolated and analyzed.Genomic DNA was extracted from the muscle,digested with restriction enzyme Sau3AI,and connected with the specific joint.The 200～900 bp DNA fragments were captured by hybridization with the biotin-probe of(CA)12.The enriched DNA fragments were ligated into a pMD-18 T-vector and then transformed into E.coli JM109 cells.Finally,the microsatellite enriched library was constructed.A total of 353 positive clones were screened from 1200 transformants,and 53 positive clones were randomly selected and then sequenced.According to sequencing results,18 pairs of primers were designed and 6 markers with high polymorphism were selected,of which the polymorphism information content(PIC) values were more than 0.5.These primers would be helpful in further analysis of genetic diversity in Tibetan chicken.The results obtained in this study show that the method of enrichment by magnetic beads could improve the efficiency of screening of microsatellite markers.