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【目的】克隆药用真菌猪苓MAPK基因并进行生物信息学分析及表达模式研究。【方法】利用5′-RACE-PCR技术从猪苓菌丝中克隆得到MAPK基因全长,利用生物信息学软件推测蛋白的理化性质、结构域;DNA Star对氨基酸进行多序列比对;用MEGA 5.0做进化关系分析;借助实时定量PCR检测基因表达模式。【结果】猪苓MAPK基因的全长c DNA为1 293 bp,其中编码区占1 161 bp,共编码386个氨基酸,推测分子量为43.872 k D,理论等电点为6.68。猪苓的MAPK有MAPK中ERK1/2类型的保守区。系统进化树结果显示猪苓MAPK蛋白属于担子菌类群。实时荧光定量PCR分析结果表明猪苓菌核形成初期,菌核中的MAPK表达量显著高于菌丝组织,随着菌核的快速生长而减少。【结论】猪苓MAPK基因Pu MAPK的分子特征为进一步研究其在猪苓菌丝形成菌核过程中的作用奠定基础。
【Objective】 Clone the MAPK gene of medicinal fungi Polyporus umbellatus for bioinformatics analysis and expression pattern. 【Method】 The full-length MAPK gene was cloned from the mycelium of Polyporus umbellatus by 5’-RACE-PCR technique. The physical and chemical properties and domains of the protein were deduced by using bioinformatics software. Multiple alignment of amino acids was performed by DNA Star. 5.0 to do evolutionary relationship analysis; real-time quantitative PCR to detect gene expression patterns. 【Result】 The results showed that the full length c DNA of MAPK gene was 1 293 bp in length, including 1 161 bp coding region encoding a total of 386 amino acids. The deduced molecular weight was 43.872 kD and the theoretical isoelectric point was 6.68. Polyporus MAPK has a conserved region of ERK1 / 2 type in MAPK. Phylogenetic tree showed that MAPK protein belonged to basidiomycetes. Real-time quantitative PCR analysis showed that the expression of MAPK in sclerotia was significantly higher than that in mycelium in the early stage of Kunxingia sclerotiorum, and decreased with the rapid growth of sclerotia. 【Conclusion】 The molecular characteristics of MAP MAPK Pu MAPK laid the foundation for further study on the role of Pu MAPK in the process of mycelium formation.