含PxIxIT模序烟曲霉KpsF基因作用初探

中国真菌学杂志　 2017, Vol. 12 　Issue (1): 8-12.

含PxIxIT模序烟曲霉KpsF基因作用初探

李佳娟¹, 李雯¹, 马彦²

1. 山西医科大学, 太原 030001;
2. 山西医科大学第二医院皮肤科, 太原 030001

收稿日期:2016-10-20 出版日期:2017-02-28 发布日期:2017-02-28
通讯作者: 马彦,E-mail:mayan197522@163.com E-mail:mayan197522@163.com
作者简介:李佳娟,女（汉族）,硕士研究生在读.E-mail：642322732@qq.com
基金资助:
国家青年科学基金（81101233），山西省青年科技研究基金（2010021036-2），山西省卫生计生委科研课题（2015043），山西省回国留学人员科研资助项目（2016-052）

The study of KpsF gene which contains PxIxIT motif in Aspergillus fumigatus

LI Jia-juan¹, LI Wen¹, MA Yan²

1. Shanxi Medical University, Taiyuan 030001, China;
2. Department of Dermatology, The Second Hospital of Shanxi Medical University, Taiyuan 030001, China

Received:2016-10-20 Online:2017-02-28 Published:2017-02-28

摘要/Abstract

摘要：

目的构建烟曲霉KpsF 基因缺陷株，初步了解KpsF基因在烟曲霉生长及与钙调磷酸酶相互作用关系。方法 PCR扩增烟曲霉KpsF基因及其上、下游各约1.0 kb的DNA片段，以pyrG为筛选标记，原生质体法构建KpsF基因缺陷株ΔKpsF；观察ΔKpsF一般生长状况；实时荧光定量PCR方法检测Ku80（对照组）和ΔKpsF（实验组）在含有不同浓度CaCl₂的液体GMM培养基中钙调磷酸酶催化亚基（CnaA）相对表达水平。结果烟曲霉ΔKpsF和Ku80径向生长差异无统计学意义（P>0.05）；表型未见明显差别；ΔKpsF在10 mmol/L CaCl₂ GMM液体培养基的CnaA相对表达量显著高于对照组Ku80（P<0.01），而100 mmol/L CaCl₂的CnaA相对表达量却没有明显变化（P>0.05）。结论 KpsF基因缺陷对烟曲霉的径向和表型生长没有明显影响，对CnaA表达量的影响与Ca²⁺浓度有关。在低浓度Ca²⁺条件下，KpsF可能参与对钙调磷酸酶的负反馈调节作用，而在高浓度的Ca²⁺条件下，这种负反馈调节作用受到抑制。

关键词: 烟曲霉, 钙调磷酸酶, KpsF基因, PxIxIT模序

Abstract:

Objective To construct the KpsF gene defect strains ΔKpsF and to explore the interaction between KpsF gene and calcineurin.Methods Amplificated the upstream and downstream of KpsF about 1.0 kb DNA fragments,the pyrG of plasmid pJW 24 was used as a biomarker when constructting the knockout plasmid.Recombinant plasmid was transformed into A.fumigatus Ku80 pyrG and got ΔKpsF.To observer the radial and phenotypic growth of Ku80 and ΔKpsF,Real-time PCR was used to determine the relative expression of calcineurin catalytic subunit (CnaA) of Ku80 (control group)and ΔKpsF (experimental group) in GMM liquid medium containing different concentration of CaCl₂.Results Compared with Ku80,the radial growth of ΔKpsF had no obvious change (P>0.05),and the phenotypic growth had no obvious difference.In GMM Liquid medium containing 10 mmol/L CaCl₂,the relative expression of CnaA of ΔKpsF strain was significantly higher than Ku80 (P<0.01).However,the relative expression of CnaA did not obviously changed (P>0.05) in GMM Liquid medium containing 100 mmol/L CaCl₂.Conclusions The KpsF had no effect in radial and phenotypic growth of A.fumigatus.Under the condition of different concentrations of Ca²⁺,the relative expression levels of CnaA gene were different between ΔKpsF and Ku80,suggesting that the KpsF might be involved in the feedback regulation of calcineurin under the condition of low concentration of CaCl₂.

Key words: Aspergillus fumigatus, Calcineurin, KpsF gene, PxIxIT motif

中图分类号:

R379.6

李佳娟, 李雯, 马彦. 含PxIxIT模序烟曲霉KpsF基因作用初探[J]. 中国真菌学杂志, 2017, 12(1): 8-12.

LI Jia-juan, LI Wen, MA Yan. The study of KpsF gene which contains PxIxIT motif in Aspergillus fumigatus[J]. Chinese Journal of Mycology, 2017, 12(1): 8-12.

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