三疣梭子蟹IAG基因结构及不同剪切体时空表达

Structural Analysis of IAG Gene and Spatiotemporal Expression Patterns of Different Spliceosomes in Portunus trituberculatus

【目的】分析三疣梭子蟹(Portunus trituberculatus)IAG(PtIAG)基因的结构及不同剪切体的时空表达模式,揭示其在性别分化中的功能及作用机制。【方法】基于NCBI数据库的甲壳动物IAG基因序列构建ML系统进化树。利用生物信息学软件对PtIAG基因的DNA结构及启动子区域进行分析。利用PCR技术对PtIAG基因进行可变剪切分析。利用qPCR技术对PtIAG基因进行时空表达分析。【结果】ML系统进化树显示,PtIAG基因与其他蟹类IAG基因的进化关系更近。PtIAG基因包含4个外显子和3个内含子,全长2615 bp。核心启动子位于起始密码子(ATG)上游2834 bp,启动子区域包括SOX-5、SOX-9、DSX等转录因子识别位点。PtIAG基因存在2种不同的转录本,其中PtIAG2为内含子保留型可变剪切转录本。qPCR结果显示,在幼体发育时期,PtIAG基因主要在溞状幼体Ⅰ期的雄蟹中表达;在成体组织中,PtIAG基因主要在促雄性腺中表达。【结论】PtIAG基因在三疣梭子蟹雄性分化及维持中具重要功能。

【Objective】To analyze the structure of IAG gene(PtIAG)and the spatial and temporal expression patterns of different spliceosomes and to reveal the function and mechanism of IAG gene in the sex differentiation of Portunus trituberculatus.【Methods】ML phylogenetic tree was constructed based on NCBI database of IAG gene sequences of crustaceans.The DNA structure and promoter region of PtIAG gene were analyzed by bioinformatics software.The variable splicing analysis of PtIAG gene was performed by PCR.The temporal and spatial expression of PtIAG gene was analyzed by qPCR.【Results】The ML phylogenetic tree showed that PtIAG gene was more closely related to IAG genes of other crabs.The length of PtIAG gene was 2615 bp,including 4 exons and 3 introns.The core promoter was located at 2834 bp upstream of the start codon(ATG),and the promoter region included transcription factor recognition sites such as SOX-5,SOX-9 and DSX.There were two different transcripts of PtIAG gene,among which PtIAG2 was an intron-retained variable shear transcript.The results of qPCR showed that PtIAG gene was mainly expressed in the male crab of zoea larva stageⅠ.In adult tissues,PtIAG was mainly expressed in androgenic gland.【Conclusion】PtIAG gene plays an important role in male differentiation and maintenance of P.trituberculatus.