Zhu Xiaohan, Sayari Mohammad, Daayf Fouad
Department of Plant Science, University of Manitoba, 222 Agriculture Building, Winnipeg, MB R3T2N2, Canada.
Agriculture and Agri-Food Canada Saskatoon Research Centre, Saskatoon, SK S7N 0X2, Canada.
Pathogens. 2021 May 23;10(6):642. doi: 10.3390/pathogens10060642.
is a hemibiotrophic pathogen responsible for great losses in dicot crop production. An gene (VDAG_03463,) identified using subtractive hybridization/cDNA-AFLP, showed higher expression levels in highly aggressive than in weakly aggressive isolates. We used a vector-free split-marker recombination method with PEG-mediated protoplast to delete the gene in This is the first instance of using this method for transformation. Only two PCR steps and one transformation step were required, markedly reducing the necessary time for gene deletion. Six mutants were identified. expressed more in the highly aggressive than in the weakly aggressive isolate in response to potato leaf and stem extracts. Its expression increased in both isolates during infection, with higher levels in the highly aggressive isolate at early infection stages. The disruption of did not influence virulence, nor did it affect total exopolygalacturonase activity in . Full genome analysis showed 8 more genes related to polygalacturonase/pectinase activity in . Transcripts of increased in the mutant in response to potato leaf extracts, compared to the wild type. The expression pattern of those eight genes showed similar trends in the mutant and in the weakly aggressive isolate in response to potato extracts, but without the increase of in the weakly aggressive isolate to leaf extracts. This indicated that the mutant of compensated by the suppression of ExoPG by activating other related gene.
是一种半活体营养型病原体，会导致双子叶作物生产遭受巨大损失。通过消减杂交/cDNA-AFLP鉴定出的一个基因（VDAG_03463）在高侵袭性菌株中的表达水平高于低侵袭性菌株。我们使用了一种无载体的分裂标记重组方法结合聚乙二醇介导的原生质体来敲除该基因，这是首次将此方法用于该菌的转化。仅需两个PCR步骤和一个转化步骤，显著减少了基因敲除所需的时间。鉴定出了六个突变体。该基因在高侵袭性菌株中对马铃薯叶片和茎提取物的响应中表达量高于低侵袭性菌株。在感染过程中，两个菌株中的该基因表达均增加，在感染早期高侵袭性菌株中的表达水平更高。该基因的敲除不影响毒力，也不影响该菌的总胞外多聚半乳糖醛酸酶活性。全基因组分析显示该菌中还有8个与多聚半乳糖醛酸酶/果胶酶活性相关的基因。与野生型相比，该基因的转录本在该菌的突变体中对马铃薯叶片提取物的响应中增加。在对马铃薯提取物的响应中，这八个基因的表达模式在该菌的突变体和低侵袭性菌株中显示出相似的趋势，但低侵袭性菌株对叶片提取物的响应中该基因没有增加。这表明该菌的该基因突变体通过激活其他相关基因来抑制外切多聚半乳糖醛酸酶从而实现补偿。
