肯尼亚拟盘多毛孢几丁质酶基因鉴定及表达分析

Identification and Expression Analysis of the Chitinases Gene Family in Pestalotiopsis kenyana

利用生物信息学方法从重寄生肯尼亚拟盘多毛孢(Pestalotiopsis kenyana)PG52菌株基因组中挖掘几丁质酶基因并分析,通过检测锈孢子诱导不同时间段基因的表达情况筛选重寄生相关几丁质酶基因。结果表明:在PG52中共鉴定到20个GH18家族和1个GH19家族几丁质酶基因,分子量38.0~177.2 kDa,理论pI值范围3.97~9.25;其中7个有信号肽,13个定位在胞外。所有PGChns基因都含有GH18家族或GH19家族保守结构域;与木霉几丁质酶序列聚类分析显示,PG52含有7个sgA、4个sgB、8个sgC、1个sgD几丁质酶基因。锈孢子诱导下共18个几丁质酶基因在转录组中被检测到表达,7个基因在诱导24 h时表达量最高,6个基因在经诱导后持续下调表达,4个基因在诱导72 h时表达量最高。对6个表达量高以及表达倍数高的基因进行RT-qPCR分析,所有基因均出现显著差异表达,并在诱导72 h时表达量最高。经诱导后上调表达的PGChns很可能在PG52菌株重寄生过程中发挥破坏锈孢子壁的作用,需要后续进一步验证。

Bioinformatics methods were used to detect and analyze chitinase genes from the genome of Pestalotiopsis kenyana PG52.The chitinase genes related to mycoparasitism were screened by detecting the expression of genes induced by aecidiospores at different time periods.Results showed the PG52 harbors 20 GH18family chitinase genes and a GH19 family gene with molecular weights of 38.0–177.2 kDa and theoretical pI of3.97–9.25,7 genes had signal peptides and 13 genes were located in extracellular.PGChns gene contained the conserved domains of the GH18 family or the GH19 family.Additionally clustering analysis with Trichoderma spp.showed that PGChns of GH18 family can be classified to 7 sgA,4 sgB,8 sgC,and 1 sgD chitinase genes.After rust spore induction,18 genes were differentially expressed,7 genes were most highly expressed at 24h induction,6 continuously down-regulated genes after induction,4 genes with most high expression after 72h induction.Subsequently,6 genes with high expression or great fold change were selected for RT-qPCR,and the results suggested the highest expression level of differently expressed genes was observed after 72h induction.The upregulated expression of PGChns after induction is likely to play a role in destroying the rust spore wall during the reparasitization of PG52 strain,while the mechanism of its reparasitization action is not yet clear.