杧果炭疽病菌 3 个果胶裂解酶基因序列特征及受漆酶基因Lac1 影响的分析

果胶裂解酶是炭疽菌在侵染寄主过程中降解寄主细胞壁的一类重要水解酶。本研究在杧果炭疽病菌中克隆获得了 3 个果胶裂解酶基因 Cgpel1、Cgpel2 和 Cgpel3，DNA 全长分别为 1 037、1 498、1 089 bp，cDNA 全长分别为 975、 1 380、978 bp，分别编码 324、459、325 个氨基酸，均含 1 个果胶裂解酶保守结构域，均有典型的信号肽，不存在跨膜结构。其二级结构中 α-螺旋分别占 16.05%、20.26%、16.92%，延伸链分别占 28.09%、21.79%、29.54%，β-转角分别占 5.86%、8.93%、7.38%，无规则卷曲分别占 50.00%、49.02%、46.15%。Cgpel1、Cgpel2 和 Cgpel3 的氨基酸序列分别与 C. tofieldiae（KZL77240.1）、草莓炭疽菌（C. gloeosporioides）（XP-007274932.1）、C. incanum（KZL84476.1）果胶裂解酶序列相似度在 93%以上。qRT-PCR 分析发现，3 个果胶裂解酶基因在整个侵染过程中均持续高效表达，但在漆酶基因 Lac1 敲除突变体中，表达均下降约 90%。可见 Cgpel1、Cgpel2 和 Cgpel3 是果胶裂解酶基因家族成员，序列差异较大，在侵染过程中起着重要的作用，且其表达受漆酶基因 Lac1 影响。

Analysis of the Characteristics of Three Pectate Lyase Gene Sequences and the Effect of Laccase Gene Lac1 on Pectin Lyase Genes

Pectate lyase is a kind of important hydrolase for the degradation of host cell wall during the process infecting host by Colletotrichum spp. In this study, Cgpel1, Cgpel2 and Cgpel3 genes were cloned from C. gloeosporioides, which causing mango anthracnose. The complete DNA sequence of Cgpel1, Cgpel2 and Cgpel3 was 1 037, 1 498 and 1 089 bp, respectively. The complete cDNA sequence was 975, 1 380 and 978 bp, encoding 324, 459 and 325 amino acids, with a specific conserved domain of pectate lyase. Cgpel1, Cgpel2 and Cgpel3 contained signal peptides and no transmembrane structure. In the secon?dary structure, there were 16.05%, 20.26% and 16.92% α-helix, 28.09%, 21.79% and 29.54% extension strand, 5.86%, 8.93% and 7.38% β-turn and 50.00%, 49.02% and 46.15% random coil. There were also great differences in the tertiary structure. Phylogenetic analysis showed that amino acid sequence of Cgpel1, Cgpel2 and Cgpel3 respectively was homologous to that of C. tofieldiae pectate lyase (KZL77240.1), C. gloeosporioides pectin lyase (XP-007274932.1) and C. incanum pectate lyase (KZL84476.1) with a similarity over 93%. Three pectate lyase genes were highly expressed continuously throughout the infection process by quantitative real-time PCR, but decreased about 90% in laccase Lac1 gene knockout mutant. The findingsindicated that Cgpel1, Cgpel2 and Cgpel3 were the members of the pectate lyase gene family, with large differences in sequence and played an important role in the process of infection. The expression were affected by laccase gene Lac1