Scning, Cloning Of Repetitive Sequences And Developing Of SCAR Markers Specific For P Genome Of Agropyron Cristatum

In this study, Agropyron cristatum (P genome) and other genome of Triticum were used to isolate specific repetitive sequence, and transformed useful RAPD marker to SCAR marker, which provides an new and effective way in detecting alien chromatin of Wheat-Agropyron additions and elucidating evolution relationship of different genomes of Triticum. The main results were obtained as follows:1. P genome plants and ABD genome plants were screened by RAPD marker, as a result, 195 RAPD polymorphism primers between them were got from 520 RAPD primers, and got 260 specific bands of P-genome. At the same time, the 195 RAPD polymorphism peimers were screened on second time, and got 78 RAPD primers with better amplification, of which 13 RAPD primers were the best. Because of the differents from species and entironment, the Agropyron tetraploid, the diploid and the hexaploid existed genetic diversity on OPA06 and OPA07.2. The genomes of part plants of Triticum were screened by RAPD markers, and P-genome specific fragments were reclaimed, from which three specific DNA repetitive sequences of P genome of Agropyron cristatum were obtained, it was OPX071036, OPX11817, and OPC051539 ,repectively. The G+C content of OPX071036, OPX11817, and OPC051539 was 37.2%, 40.8%, and 41.3%, repectively. Homologous comparison with the DNA sequences registered in GenBank (GenBank + EMBL + PDB) indicated that it were the new P-genome specific DNA repetitive sequence of Agropyron.3. Three pairs of SC-OPX07, SC-OPX11, and SC-OPC05 specific primers were designed according to the sequence of OPX071036, OPX11817, and OPC051539, repectively. The results of PCR indicated that OPX071036 and OPX11817 could be used to distinguish P genome from genome ABD, C, D, E, AG, H, I, M, Ns, R, S, StPY, U, V, and St, while OPC05815 could distinguish P genome from genomes ABD, C, D, E, AG, I, M, R, S, U, and V.4. OPX071036, OPX11817, and OPC051539 was used as probes, repectively. southern hybridization showed that they all had strong distribution in the P genome, but no hybridization signals detected in other genomes, which consistent with the results of PCR amplification. Furthermore, the tetraploid Agropyron (Z2044) was studied by FISH using pAX07, pAX11, and pAC05 as probes, repectively. the results showed that they all distributed on the whole chromosomes of Z2044, but it distribute differently to each chromosome. These results above indicated that OPX071036, OPX11817, and OPC051539 were the specific DNA repetitive sequence of P genome, the three specific probes and the corresponding SCAR markers were all used to detect the alien chromatin of Wheat-Agropyron additions.