Identification of pathogenicity mutants of the rice blast fungus Magnaporthe grisea by insertional mutagenesis
Identification of pathogenicity mutants of the rice blast fungus Magnaporthe grisea by insertional mutagenesis
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DOI：
10.1094/mpmi.1999.12.2.129
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发表时间：
1999-02-01
影响因子：
3.5
通讯作者：
Talbot, NJ
中科院分区：
文献类型：
--
作者：
Balhadère, PV;Foster, AJ;Talbot, NJ
采用限制性酶介导的DNA整合（REMI）突变来鉴定稻瘟菌致病性受损的突变体。评估了三种REMI方案，并确定了REMI的频率。一个REMI文库3，527 M。在三种遗传背景中产生稻瘟病菌转化体，用大麦切叶测定筛选1，150个转化体的致病性缺陷。五个突变体进行了鉴定和表征。两个突变体（2029和2050）的附着胞功能受损。另外两个突变体，125和130，改变了分生孢子形态，分生孢子发生，附着胞功能。突变体130也是一个蛋氨酸营养缺陷型和蛋氨酸营养缺陷型共分离的致病性降低。另外一个突变体，80，表现出降低对稻瘟病易感水稻品种的致病性，但完全致病大麦。突变体80的致病性降低与孢子萌发和附着胞发育延迟有关。遗传分析表明，每个突变体的单基因分离，但只有两个突变与潮霉素抗性标记共分离。突变体2029、2050、125、130和80中的遗传位点分别被称为PDE 1、PDE 2、IGD 1、MET 1和GDE 1。pde1和pde2与cpkA是非等位基因，cpkA是环腺苷酸（cAMP）依赖性蛋白激酶A催化亚基中的突变，具有非常相似的表型。结果表明，实用的REMI研究真菌致病性，但也强调了严格的遗传和表型分析的要求。
Restriction enzyme-mediated DNA integration (REMI) mutagenesis was used to identify mutants of Magnaporthe grisea impaired in pathogenicity. Three REMI protocols were evaluated and the frequency of REMIs determined. An REMI library of 3,527 M. grisea transformants was generated in three genetic backgrounds, and 1,150 transformants were screened for defects in pathogenicity with a barley cut leaf assay. Five mutants were identified and characterized. Two mutants (2029 and 2050) were impaired in appressorium function. Two other mutants, 125 and 130, were altered in conidial morphology, conidiogenesis, and appressorium function. Mutant 130 was also a methionine auxotroph and methionine auxotrophy cosegregated with the reduction in pathogenicity. An additional mutant, 80, showed reduced pathogenicity on blast-susceptible rice cultivars but was fully pathogenic on barley. The reduction of pathogenicity in mutant 80 was associated with a delay in conidial germination and appressorium development. Genetic analysis suggested single-gene segregation for each mutant, but only two of the mutations co-segregated with the hygromycin resistance marker. The genetic loci in mutants 2029, 2050, 125, 130, and 80 were termed PDE1, PDE2, IGD1, MET1, and GDE1, respectively. pde1 and pde2 were non-allelic to cpkA, a mutation in the catalytic subunit of cyclic AMP (cAMP)-dependent protein kinase A with a very similar phenotype. The results indicate the utility of REMI for studying fungal pathogenicity, but also highlight the requirement for rigorous genetic and phenotypic analysis.