Isolation And Characterization Of Resistance Gene From Wild Rice Genomic Library

The genome of the wild rice, which contains large amounts of resistance genes, is an important gene resource in rice breeding. Because of the complex genetic background of the wild rice, the useful gene is difficult to obtain by conventional gene cloning methods. The large-fragments insert genomic library provides a new platform for developing the useful genes of wild rice. But there are more than 10,000 clones in the genomic library, it would cost a huge labor and quite a long time to transfer every clone to the rice cultivar. In order to explore the useful germplasm of the wild rice in a short period, the following work has been done:(1) A hybridization system of screening resistance gene from genomic library was established after optimizing the hybridization conditions. The system was tested by a positive example and the results indicated that the method of screening wild rice genomic library was available.(2) According to the conserved sequences of the existing resistance genes, two kinds of probes were synthesized. The genomic libraries of Dongxiang wild rice and Xiaoli wild rice were screened by the colony hybridization. 49 positive clones were obtained. The positive clones have been transferred into the rice cultivar.(3) Some positive clones were identified by polymerase chain reaction, the purified fragments were TA cloned and sequenced. Five sequences were obtained. Sequence analysis indicated that the five sequences were homologous with parts of the cloned resistance gene sequences. So the obtained sequences could be parts of the candidate resistance genes.The establishment of the system supplies us a new idea of developing useful genes from wild rice genomic library. The number of clones, which would be used to transfer into rice cultivar, has been reduced into 49 from more than 10,000. The confirmed positive clones will be used to construct the subclone and have a further sequence.