Molecular And Cytogenetic Identification Of Wheat-dasypyrum Breviaristatum Introgression Lines And Development Of New Specific Markers

As a wild relative of wheat species,Dasypyrum is displayed drought tolerance and diseases resistance, as well as extensive range of its growth. Especially, many excellent disease resistant genes of Dasypyrum villosum have imported in wheat background, which greatly improves the disease resistance of wheat, Therefore it serves as excellent source for the genetic improvement of wheat. But the current studies on the research of the Dasypyrum beviaristatum is rather limited, and its excellent genes are not transferred to wheat background, and which limiting the application of its good properties. By using Sichuan wheat hybridized and backcrossed with wheat-Dasypyrum breviaristatum partial amphiploid, we created a series of wheat-D. breviaristatum introgression lines, which enrich application of the excellent genes of D. breviaristatum in wheat breeding.In this study, total genomic DNA of D. breviaristatum was used as probe for genomic in situ hybridization(GISH) analysis of wheat-Dasypyrum breviaristatum introgression lines, the results indicates the two whole arms chromosome segments of D. breviaristatum were introgressed in wheat background. Moreover, the Oligo based probes, Oligo-pSc119.2, Oligo-pTa535 were used as probes in fluorescence in situ hybridization(FISH) analysis of the wheat-D. breviaristatum introgression lines. The FISH result indicates a pair of 5AS chromosome of D2146, D2147, D2148, D2149 translocated to the whole arm of chromosome of D. breviaristatum, and whole arm of chromosome of D. breviaristatum can be 1VbL, 1VbS, 2 VbL, 3 VbL, or 5 VbL. In order to identify the alien chromosome segments, we screened 145 pairs of PLUG primer, 256 pairs of EST primer and 56 pairs of COS primer. The PCR amplification techniques combined with restriction enzyme digestion analysis were used,.We found the 9 pairs of primer located on chromosome of homologous group 5 of wheat or rice or barleyTNAC1485, TNAC1554, TNAC1567, TNAC16181, CINAU157 a, CINAU446, CINAU448, CINAU455, COS93 can specific amplify in D. breviaristatum, D2146, D2147, D2148, D2149 but not in Chinese Spring, which indicated that alien chromosome segments belong to homologous group 5, which means the alien chromosome segments were 5VbL or 5VbS. So the FISH and PCR result indicated D2146, D2147, D2148, D2149 are wheat-D. breviaristatum 5AS·5VbL translocation line. The 8 pairs of primer- TNAC1485, TNAC1554, TNAC1567, TNAC16181, CINAU157 a, CINAU446, CINAU448, COS93 can be used as new moleculer marker, which can fast and accurately identify the chromosome segment of D. breviaristatum in wheat background and confirm its homology.We also found that D2146, D2147, D2148, D2149 showed late heading, so the 11 pairs of primer based on gene Vrn-1, Vrn-2, Vrn-3 controlling vernalization were detected. The results showed that VrnB primers can amplify in Chinese Spring but D2146, D2147, D2148, D2149. However VrnB1, VrnB3 primer can amplify in D2146, D2147, D2148, D2149, but Chinese spring not. So we can speculate that the VrnB1, VrnB3 gene maybe control late heading.