稻瘟病新抗性基因Pita2候选基因的表达载体构建

Construction of Expression Vector for Candidate Genes of New Rice Blast Resistance Gene Pita2

为了验证稻瘟病新抗性基因Pita2的功能,利用长片段PCR技术从基因的供体品种PiNo.4中扩增了2个候选基因Pita2-1,Pita2-2,长度分别为6.1 kb和16.5 kb,电泳回收目的片段,然后利用双酶切分别克隆到植物表达载体pCAMBIA1300。对阳性克隆通过菌落PCR、酶切鉴定和测序分析,已成功地获得了2个候选基因的重组阳性克隆,为进一步研究Pita2基因的功能奠定了基础。

Rice blast is one of the most devastating diseases of rice,and a serious threat to rice production.Exploring new rice blast resistance genes for breeding disease resistance strains is an important strategy.To study the function of two candidate genes Pita2-1 and Pita2-2,long-range PCR（LR-PCR） was employed to amplify the candidate genes from genomic DNA of the resistant cultivar PiNo.4.The appropriate products of Pita2-1（6.1 kb）and Pita2-2（16.5 kb） were purified and then ligated into the binary vector pCAMBIA1300 respectively.The positive clones were confirmed by colony PCR,restriction endonuclease digestion and sequencing,thus laid a foundation for dissecting the function of gene Pita2.