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野生蕉是香蕉遗传改良的天然基因库。为了分离野生蕉中的抗病基因类似序列,根据核苷酸结合位点(nucleotide binding site,NBS)和丝氨酸/苏氨酸蛋白激酶域设计简并性引物,以经过鉴定的抗香蕉枯萎病4号生理小种的野生蕉(Musa acuminata)叶片cDNA为模板进行PCR扩增。经过对扩增产物进行克隆和测序,获得了6个500 bp左右的RGAs片段。其中有2个RGA(WNB1和WNB2)具有NB-ARC保守结构域特征,并且WNB1具有连续的ORF。其余4个RGAs(WST1、WST2、WST3和WST4)均具有丝氨酸,苏氨酸蛋白激酶域特征,且WST3编码的氨基酸序列与水稻抗叶斑病基因Xa21同源性很高。用半定量PCR分析枯萎病菌诱导后野生蕉叶片中RGAs的表达情况,结果表明WNB1和WST3受枯萎病菌诱导后表达量增强,这表明WNB1和WST3的表达可能与香蕉枯萎病抗性相关。
Wild banana is a natural gene bank for genetic modification of bananas. In order to isolate similar sequences of the resistance genes in wild banana, degenerate primers were designed based on the nucleotide binding site (NBS) and serine / threonine protein kinase domains, and identified anti-banana wilt disease 4 Number of physiological races wild Musa (Musa acuminata) leaf cDNA as a template for PCR amplification. After cloning and sequencing the amplified products, 6 RGAs fragments of about 500 bp were obtained. Among them, 2 RGA (WNB1 and WNB2) have NB-ARC conserved domain features, and WNB1 has a continuous ORF. The remaining four RGAs (WST1, WST2, WST3 and WST4) all have serine and threonine protein kinase domains, and the amino acid sequence encoded by WST3 has high homology with the leaf spot resistance gene Xa21 in rice. Semi-quantitative PCR analysis of RGAs expression in wild banana leaves induced by Fusarium oxysporum f. Sp. Showed that the expression of WNB1 and WST3 was enhanced by Fusarium oxysporum f. Sp. Infection, suggesting that the expression of WNB1 and WST3 may be related to the resistance of banana wilt disease.