摘要
为了寻找并克隆凡纳滨对虾(Litopenaeus vannamei)G蛋白α亚基,为研究对虾的生理调控提供理论基础,通过简并PCR和RACE反应获得目的基因的全长cDNA序列;用Blast,DNAstar和Genedoc软件对所得序列进行分析,确定所得序列为凡纳滨对虾G蛋白Gαq基因,命名为pvGqα。将该基因的编码序列克隆到表达载体,通过免疫共沉淀实验和胞内Ca2+,IP3浓度的测定鉴定该Gαq亚基的功能,发现该亚基的序列和功能与其他Gαq家族成员具有高度保守性。用Western blotting分析发现pvGαq在对虾身体各部位存在普遍分布,尤其在脑神经、眼柄、腮和颚片中有大量表达,在嗅觉器官触角也有适量分布。说明了pvGαq在对虾生命活动中的重要性,为研究对虾的生理调控奠定了理论基础。
In this study, we aimed to identify new functional genes encoding G protein alpha subunits from Litopenaeus vannamei and elucidate the critical roles that Gα subunits assume in a signal transduction process of shrimp. Degenerate PCR and RACE techniques were employed to achieve the full length of Gα subunit in L. vannamei. Co-immunoprecipitation analysis and measurement of second messengers of Ca^2＋ and IP3 in HEK 293 cells overexpressed with this new Gα subunit were used to identify the basic functions. Western blotting analysis was employed to detect the distribution of the Gα subunit. In the results, the predicted encoding protein of the novel cDNA sequence shared high similarity to Gαq from other organisms, and was thus termed pvGαq, pvGαq was highly conserved in both primary structures and basic functions. The expression pattern revealed that pvGαq was widely expressed in many shrimp tissues, especially enriched in brain and eyes, it also obviously existed in gills, maxillas and antennae. These results indicated that pvGαq may play critical roles in sensory responses and nerve system of shrimp L. vannamei, which provided a basis for the research of physiological regulation of shrimp.
出处
《海洋科学》
CAS
CSCD
北大核心
2008年第8期64-69,共6页
Marine Sciences
基金
国家863计划资助项目(2002AA629060)
