草菇MAPKKK同源基因的克隆及序列分析

Cloning and Sequence Analysis of Mitogen-activated Protein Kinase Kinase Kinase Homolog from Volvariella volvacea (VV-MAPKKK)

摘要:

根据担子菌丝裂原活化蛋白质激酶激酶激酶(MAPKKK)蛋白的保守序列设计两对简并引物, 通过巢式简并PCR方法获得草菇VV-MAPKKK基因中的保守片段, 然后通过和草菇基因组信息比对, 获得了VV-MAPKKK基因全长序列。VV-MAPKKK基因长度为4434 bp, 包含4个内含子, 编码1405个氨基酸残基, 推定的氨基酸序列与新型隐球菌(Cryptococcus neoformans)、巴西芽生菌(Paracoccidioides brasiliensis)和异旋孢腔菌(Cochliobolus heterostrophus)的MAPKKK同源蛋白相似性分别为58%、57%和56%。对VV-MAPKKK蛋白的系统发生学分析的结果表明, VV-MAPKKK与担子菌中的Hog信号传导途径的MAPKKK同源蛋白聚在同一进化支上, 这些数据都支持所获得的VV-MAPKKK为Hog-MAPKKK蛋白在草菇中的同源物的推定。

关键词:草菇, 丝裂原活化蛋白质激酶激酶激酶(MAPKKK), 克隆, 序列分析

Abstract:

Two pairs of degenerate primers were designed according to the conserved amino acid sequences of several mitogen-activated protein kinase kinase kinases (MAPKKK) from basidiomycetes. Based on the conserved DNA sequence, which was obtained by nested degenerate PCR, a whole DNA sequence of MAPKKK of Volvariella volvacea (VV-MAPKKK) was obtained by blasting with Volvariella volvacea genome. VV-MAPKKK gene, containing 4 introns and codeing for a 1405 aa protein (VV-MAPKKK), is 4434 bp in length. The similarity of VV-MAPKKK compared with Hog-MAPKKK protein from Cryptococcus neoformans, Paracoccidioides brasiliensis and Cochliobolus heterostrophus is 58%, 57% and 56%, respectively. Phylogenetic analysis showed that VV-MAPKKK and Hog-MAPKKK protein homolog belong to the same clade. These data supported that VV-MAPKKK protein is a Hog-MAPKKK protein homolog from Volvariella volvacea.

Key words:Volvariella volvacea, Mitogen-activated protein kinase kinase kinase (MAPKKK), Clone, Sequence analysis