生物
突变体
基因沉默
基因
RNA干扰
木聚糖酶
遗传学
毒力
细胞生物学
酶
核糖核酸
生物化学
作者
Quoc Son Nguyen,Kenji Itoh,Vu Van Ba,Yukio Tosa,Hitoshi Nakayashiki
标识
DOI:10.1111/j.1365-2958.2011.07746.x
摘要
Due to functional redundancy, it is often difficult to genetically analyse the biological function of fungal cell wall-degrading enzymes that belong to a gene family. To overcome this difficulty, we used RNAi to knock-down (KD) multiple xylanase genes to elucidate their roles in the pathogenicity of the blast fungus, Magnaporthe oryzae. To obtain the maximum average efficiency of gene silencing for the xylanase genes, we used the 'building blocks method', in which a 40 bp sequence was chosen from an endoxylanase gene, and 10 such sequences from 10 endoxylanases were combined to make an artificial RNAi trigger by synthetic DNA. Quantitative RT-PCR analysis revealed that the transcript levels of all the expressed xylanase genes were significantly reduced in KD mutants with the artificial RNAi trigger. Even though the KD mutants did not completely lose their pathogenicity to host plants, the number of lesions, rate of penetration and extent of infected cells were all reduced in KD mutant-infected leaves. The degree of pathogenicity reduction was associated with the silencing levels of xylanase mRNA and enzymatic activity in the KD mutants. Cytological analysis indicated that xylanases play significant roles in both vertical penetration and horizontal expansion of M. oryzae in infected plants.
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