渐狭蜡蚧菌的鉴定及其几丁质酶基因<em>LACHI1</em>的克隆

doi:10.7668/hbnxb.2014.04.011

摘要/Abstract

摘要： 克隆渐狭蜡蚧菌几丁质酶基因，为进一步明确该菌产生的几丁质酶对定居性根结类和孢囊类线虫卵孵化抑制作用提供理论依据。采用分离自黑龙江大豆孢囊线虫孢囊寄生真菌CGMCC5328，通过形态学特征及ITS序列比较分析，鉴定该菌株为渐狭蜡蚧菌。通过简并引物设计和RACE技术，克隆渐狭蜡蚧菌中几丁质酶基因，利用生物学软件分析该基因序列及其编码的氨基酸序列。首次从渐狭蜡蚧菌中克隆得到一个几丁质酶基因 LACHI1，该基因DNA序列长1 743 bp，含3个内含子，包含1 272 bp开放阅读框，编码423个氨基酸，理论分子量45.9 kDa，等电点5.90。 LACHI1 基因编码的几丁质酶，属于糖基水解酶18家族几丁质酶。同源性比对表明该菌产生的几丁质酶与昆虫寄生真菌和食线虫真菌产生的几丁质酶有很高的同源性。

关键词: 渐狭蜡蚧菌, 几丁质酶基因, 克隆, 系统发育分析

Abstract: To clone chitinolytic genes from Lecanicillium attenuatum and provide theoretical basis for the inhibition of the chitinases on egg-hatching of root knot and cyst nematode.In this study,an nematode egg-parasitic fungus CGMCC5328 was isolated from Heterodera glycines infecting soybean in Heilongjiang Province.Based on morphological characters and molecular analysis of ITS-rDNA,the strain was identified as L.attenuatum.The chitinase gene LACHI1 from L.attenuatum was cloned using the degenerate PCR primers and RACE techniques.The analysis of the gene LACHI1 and its amino acid sequence was by biology software.We have cloned a chitinase gene LACHI1 from L.attenuatum for the first time.The gene is 1 743 bp in length and contains three putative introns.The ORF of LACHI1 is 1 272 bp in size with encoding protein of 423 aa,molecular mass of 45.9 kDa and pI of 5.90.The chitinase deduced by LACHI1 belongs to glycosyl hydrolase family 18 chitinase.Comparison of the chitinase amino acid sequence with other chitinases from entomopathogenic fungi and nematophagous fungi revealed that the enzymes were highly similar.

Key words: Lecanicillium attenuatum, Chitinase gene, Cloning, Phylogenetic analysis

中图分类号:

赵洋, 陈德鑫, 王凤龙, 黄化刚, 武侠. 渐狭蜡蚧菌的鉴定及其几丁质酶基因LACHI1的克隆[J]. 华北农学报, 2014, 29(4): 63-70. doi: 10.7668/hbnxb.2014.04.011.

ZHAO Yang, CHEN De-xin, WANG Feng-long, HUANG Hua-gang, WU Xia. Identification of Fungus Lecanicillium attenuatum and Cloning of Its Chitinase Gene LACHI1[J]. journal1, 2014, 29(4): 63-70. doi: 10.7668/hbnxb.2014.04.011.

http://www.hbnxb.net/CN/Y2014/V29/I4/63

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渐狭蜡蚧菌的鉴定及其几丁质酶基因LACHI1的克隆

Identification of Fungus Lecanicillium attenuatum and Cloning of Its Chitinase Gene LACHI1

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渐狭蜡蚧菌的鉴定及其几丁质酶基因LACHI1的克隆

Identification of Fungus Lecanicillium attenuatum and Cloning of Its Chitinase Gene LACHI1

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