产壳聚糖酶内生真菌的筛选、鉴定及酶活力初步研究

金秋珠<sup>1; 2; 3</sup>; 颜桢灵<sup>1; 2; 3</sup>; 黄植清<sup>1; 2; 3</sup>; 袁海莹<sup>1; 2; 3</sup>; 刘广华<sup>1; 2; 3</sup>; 农小霞<sup>1; 2; 3</sup>; 李鑫<sup>1; 2; 3</sup>; 骆海玉<sup>1; 2; 3*</sup>

引用本文:	金秋珠, 颜桢灵, 黄植清, 袁海莹, 刘广华, 农小霞, 李鑫, 骆海玉.产壳聚糖酶内生真菌的筛选、鉴定及酶活力初步研究[J].广西植物,2020,40(9):1332-1340.[点击复制]
	JIN Qiuzhu, YAN Zhenling, HUANG Zhiqing, YUAN Haiying, LIU Guanghua, NONG Xiaoxia, LI Xin, LUO Haiyu.Screening and identification of chitosanase-producing endophytic fungi as well as their chitosanase activities[J].Guihaia,2020,40(9):1332-1340.[点击复制]

【打印本页】【下载PDF全文】【查看/发表评论】【下载PDF阅读器】【关闭】

←前一篇|后一篇→

过刊浏览高级检索

本文已被：浏览 5813次下载 1878次	码上扫一扫！
分享到：微信更多字体:加大+\|默认\|缩小-
产壳聚糖酶内生真菌的筛选、鉴定及酶活力初步研究
金秋珠^1,2,3, 颜桢灵^1,2,3, 黄植清^1,2,3, 袁海莹^1,2,3, 刘广华^1,2,3, 农小霞^1,2,3, 李鑫^1,2,3, 骆海玉^1,2,3*
1. 珍稀濒危动植物生态与环境保护教育部重点实验室, 广西桂林541006;2. 广西珍稀濒危动物生态学重点实验室, 广西桂林 541006;3. 广西师范大学生命科学学院, 广西桂林 541006

摘要:

植物内生真菌是挖掘不同类型壳聚糖酶及发现新酶的资源宝库。该研究从122株柑橘和血散薯内生真菌中筛选能产生壳聚糖酶的菌株,对其进行鉴定,初步研究酶活力影响因素,为后期其酶学性质及产壳聚糖酶内生真菌与宿主植物病害防御互作关系的研究奠定基础。通过透明圈法初筛结合液体发酵法进行复筛,得到2株可产生壳聚糖酶的内生真菌Stdif9和Stdif9-4,并发现Stdif9-4最高酶活力(0.968 U·mL^-1)显著高于Stdif9(0.780 U·mL^-1)。采用形态学和分子生物学结合的方法将菌株Stdif9-4鉴定为青霉属菌株,即Penicillium sp. Stdif9-4。通过DNS试剂法初步研究影响该菌株产壳聚糖酶活力的因素,发现不同培养时间对菌株壳聚糖酶活力具有显著影响,在培养96 h时,壳聚糖酶活力达到最大值。9种金属离子对菌株的酶活力具有不同影响,其中Mn²⁺和Ca²⁺对壳聚糖酶活力具有明显的激活作用; Ag⁺、Zn²⁺、Cd²⁺、Ba²⁺和Fe³⁺对壳聚糖酶活力具有不同程度的抑制作用,并且Ag⁺的抑制作用最为显著; K⁺和Na⁺对壳聚糖酶活力无显著影响。不同培养代数菌株产酶活力无显著差异,说明其产酶活力稳定。

关键词: 植物内生真菌, 壳聚糖酶, 酶活力, 青霉属, 血散薯

DOI：10.11931/guihaia.gxzw201906015

分类号:Q939.99

文章编号:1000-3142(2020)09-1332-09

基金项目:广西自然科学基金(2016GXNSFBA380049,2018GXNSFAA281013); 广西中青年教师基础能力提升项目(2017KY0080); 广西珍稀濒危动物生态学重点实验室研究基金(GKN. 17-A-02-02); 广西师范大学生态学博士点建设经费资助项目(EDPC 2018002); 2019年广西壮族自治区大学生创新创业训练计划项目(201910602244)[Supported by Natural Science Foundation of Guangxi, China(2016GXNSFBA380049, 2018GXNSFAA281013); Youth and Middle-Aged Basic Ability Promotion Program(2017KY0080); Guangxi Key Laboratory of Rare and Endangered Animal Ecology(GKN. 17-A-02-02); Ecological Doctoral Program Construction of Guangxi Normal University(EDPC 2018002); Innovation and Entrepreneurship Training Program for College Students of Guangxi(201910602244)]。

Screening and identification of chitosanase-producing endophytic fungi as well as their chitosanase activities

JIN Qiuzhu^1,2,3, YAN Zhenling^1,2,3, HUANG Zhiqing^1,2,3, YUAN Haiying^1,2,3, LIU Guanghua^1,2,3, NONG Xiaoxia^1,2,3, LI Xin^1,2,3, LUO Haiyu^1,2,3*

1. Key Laboratory of Ecology of Rare and Endangered Species and Environmental Protection, Ministry of Education, Guilin 541006, Guangxi, China;2. Guangxi Key Laboratory of Rare and Endangered Animal Ecology, Guilin 541006, Guangxi, China;3. College of Life Sciences, Guangxi Normal University, Guilin 541006, Guangxi, China 1. Key Laboratory of Ecology of Rare and Endangered Species and Environmental Protection, Ministry of Education, Guilin 541006, Guangxi, China; 2. Guangxi Key Laboratory of Rare and Endangered Animal Ecology, Guilin 541006, Guangxi, China; 3. College of Life Sciences, Guangxi Normal University, Guilin 541006, Guangxi, China

Abstract:

Endophytic fungi are good sources for exploring high diversity of chitosanase and discovering new enzymes. In this study, chitosanase-producing strains were screened and identified from 122 endophytic fungi associated with citrus and Stephania dielsiana, and the factors affecting the activity of chitosanase were preliminarily studied, in order to provide an experiment basis for further study on the enzymatic properties and the interaction between chitosanase-producing endophytic fungi and host plant resistance to diseases. Two endophytic fungi Stdif9 and Stdif9-4 were found to produce chitosanase by transparent circle method and liquid fermentation method. The highest activity of Stdif9-4(0.968 U·mL^-1)was significantly higher than that of Stdif9(0.780 U·mL^-1). Stdif9-4 was identified as Penicillium sp. Stdif9-4 by morphological method and sequencing analysis of ITS gene. The factors affecting on the chitosanase activity were studied by DNS reagent method. The results showed that the chitosanase activity reached maximum at 96 h of cultivation. In addition, among the nine metal ions, Mn²⁺ and Ca²⁺ had obvious activation effect on chitosanase activity. Ag⁺, Zn²⁺, Cd²⁺, Ba²⁺ and Fe³⁺ had different inhibitory effects on chitosanase activity, and Ag⁺ showed more significant inhibitory effect, while K⁺ and Na⁺ did not show significant effect on enzyme activity. There was no significant differences in enzyme activity between different cultured generations, which indicate that the enzyme activity is stable.

Key words: plant endophytic fungi, chitosanase, enzyme activity, Penicillium, Stephania dielsiana