中国卤虫(Artemia sinica)早期胚胎发育相关基因表达序列标签(ESTs)分析

THE EXPRESSED SEQUENCE TAGS OF ARTEMIA SINICA GENE IN EARLY EMBRYOS DEVELOPMENT

以mRNA差异显示技术分析了中国两性生殖卤虫早期胚胎发育相关基因的表达序列标签(ESTs),共回收了2000条差异条带并进行了克隆、测序,在GenBank/DDBJ/EMBL数据库中用BLASTX和BLASTN软件分别与NCBI上的非冗余核酸序列数据库进行同源性比对,得到分值较高、长度在100bp—2kb以内的ESTs序列133条。所发现的133条ESTs的生物学功能分类为:能量代谢占24%、细胞生长、分裂相关基因占23%、RNA相关占17%、体节形成占12%、蛋白质合成占3%、转录因子占3%、离子转运占5%、细胞信号转导占4%、未知占9%。对所发现的ESTs进行了RT-PCR测定,排除假阳性能够提供足够数量的信息用于获取其全长cDNA,进而获得其全长基因序列以用于功能研究。另外,研究发现了包括小热休克蛋白基因家族、Hox基因家族的abdA和Dfd基因、A-trh基因(排盐器官及盐调节基因)、HIF(缺氧诱导因子)、CK-M2(肌肉型磷酸激酶基因)和GRP基因(富甘氨酸蛋白基因)等20个具有明显功能的相关基因的ESTs和完整基因序列和开放阅读框(ORF)。经初步推断这些ESTs和基因的功能大多为与卤虫发育的生理、生化相关的基因有关。所研究的133条ESTs基本代表了卤虫早期胚胎发育相关功能基因的表达谱。

The expressed sequence tags （ESTs） of Artemia sinica genes in early embryonic development were analyzed with mRNA differential display technique. 2000 differential fragments were isolated, cloned, and sequenced, for which the sequences were compared by BLASTX by BLASTN software against the unredundance nucleotide and protein sequences in the Database of GenBank/DDBJ/EMBL. Among them 133 ESTs had higher score and ranged from 100bp--2kb. Twenty full-length sequences were sequenced completely having open reading frames （ORFs）, such as small heat shock gene family, abdA gene and Dfd gene of Hox gene family, A-trh （salt gland development gene） GRP gene （glycine-rich protein gene）, HIF （Hypoxia-inducible factor）, CK-M2 （muscle-type creatine kinase）, and so on. The biological allocation of the 133 ESTs in biological functions included 24% in energy metabolism, 23% for cell growth and division, 17% for RNA related gene, 12% for body segment formation, 3% in protein synthesis, 3% for transcription factor, 4% for cell signal transduction, 5% for ionic transport, and 9% for other unknown roles. The expression of 21 genes were analyzed by RT-PCR, from which the ESTs/gene was deduced representing the expression of Artemia sinica genes relating early embryonic development.