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Han X.,Central South University | Han X.,China National Hybrid R and nter | Han X.,Hunan Longping Gene Co. | Li L.,Central South University | And 6 more authors.
African Journal of Biotechnology | Year: 2011

A strategy was described for the isolation of disease resistance genes from Oryza minuta by integrating the techniques of transformation-competent genomic library, RecA-mediated magnetic bead enrichment library and candidate disease resistance gene cloning. The principal advantages of this method were: simple, rapid and suitable. In this research, a transformation-competent genomic library with a volume of 2.68 × 10 5 clones was constructed for O. minuta; an enrichment library of disease resistance genes was further constructed with a volume of 4992 clones, from which 26 positive clones were screened by colony in situ hybridization. The end-clone sequencing of 13 representative positive clones showed that 6 clones were well matched with cloned disease resistance genes or located near the existing disease resistance genes. Full sequencing of a clone revealed a gene similar to a putative brassinosteroid LRR receptor kinase in japonica rice; the protein structure analysis suggested that it may be a disease resistance gene or functionally involved in a signal transduction pathway. These results indicate these clones may include new R genes and the strategy is feasible to clone new R genes from wild rice species. © 2011 Academic Journals. Source


Han X.,Central South University | Han X.,China National Hybrid R and nter | Zuo J.,Central South University | Zuo J.,China National Hybrid R and nter | And 10 more authors.
Advanced Science Letters | Year: 2011

A strategy is described here for isolation of candidate disease resistance (R) genes from Dongxiang common wild rice (O. rufipogon Griff.). This method integrates the techniques of candidate gene analogs cloning, the construction of wild rice transformation competent genomic library and the R gene enrichment library, the colony in-situ hybridization experiment, and the sequence analysis of function genes. In this research, a transformation competent genomic library with a volume of 1.58×10 5 clones was constructed for Dongxiang common wild rice and an enrichment library of R genes was further constructed with a volume of 3072 clones, from which 17 positive clones were isolated. The end-clone sequencing of 9 representative positive clones reveals that 6 can be located at or near to the existed R genes or the predicted R genes. Sequences analysis of 6 clones suggests that 5 contain the conserved domain of R genes, indicating the strategy is feasible and simple to clone new R genes from wild rice species. © 2011 American Scientific Publishers. Source

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