Scott-Craig J S, Panaccione D G, Cervone F, Walton J D
Department of Energy Plant Research Laboratory, Michigan State University, East Lansing 48824-1312.
Plant Cell. 1990 Dec;2(12):1191-200. doi: 10.1105/tpc.2.12.1191.
A gene (PGN1) encoding extracellular endopolygalacturonase was isolated from the fungal maize pathogen Cochliobolus carbonum race 1. A probe was synthesized by polymerase chain reaction using oligonucleotides based on the endopolygalacturonase amino acid sequence. Genomic and cDNA copies of the gene were isolated and sequenced. The corresponding mRNA was present in C. carbonum grown on pectin but not on sucrose as carbon source. The single copy of PGN1 in C. carbonum was disrupted by homologous integration of a plasmid containing an internal fragment of the gene. Polygalacturonase activity in one transformant chosen for further analysis was 10% or 35% of the wild-type activity based on viscometric or reducing sugar assays, respectively. End product analysis indicated that the residual activity in the mutant was due to an exopolygalacturonase. Pathogenicity on maize of the mutant lacking endopolygalacturonase activity was qualitatively indistinguishable from the wild-type strain, indicating that in this disease interaction endopolygalacturonase is not required. Either pectin degradation is not critical to this interaction or exopolygalacturonase alone is sufficient.
从玉米真菌病原体玉米小斑病菌1号小种（Cochliobolus carbonum race 1）中分离出一个编码胞外内切多聚半乳糖醛酸酶的基因（PGN1）。利用基于内切多聚半乳糖醛酸酶氨基酸序列的寡核苷酸通过聚合酶链反应合成了一个探针。分离并测定了该基因的基因组和cDNA拷贝的序列。相应的mRNA存在于以果胶为碳源生长的玉米小斑病菌中，而不存在于以蔗糖为碳源生长的菌株中。通过含有该基因内部片段的质粒的同源整合破坏了玉米小斑病菌中PGN1的单拷贝。基于粘度测定或还原糖测定，选择用于进一步分析的一个转化体中的多聚半乳糖醛酸酶活性分别为野生型活性的10%或35%。终产物分析表明，突变体中的残留活性归因于外切多聚半乳糖醛酸酶。缺乏内切多聚半乳糖醛酸酶活性的突变体对玉米的致病性在定性上与野生型菌株没有区别，这表明在这种病害互作中不需要内切多聚半乳糖醛酸酶。要么果胶降解对这种互作不重要，要么仅外切多聚半乳糖醛酸酶就足够了。
