日本血吸虫期别差异表达基因文库的构建及分析
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国家高技术研究发展计划(863)资助项目(2004AA2Z3510,2001AA215151), 上海市重大科技公关项目(03DZ19231).


Construction and Analysis of Differentially Expressed Gene cDNA Library From Schistosoma japonicum in Different Development Stage
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This work was supported by grants from Hi-Tech Research and Development Program of China (2004AAZ3510, 2001AA215151) and Ministry of Major Science & Technology of Shanghai (03DZ19231).

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    摘要:

    为从期别差异表达基因分析入手研究血吸虫的生长发育机制,应用抑制性消减杂交 (suppressed subtractive hybridization , SSH) 技术首次构建了日本血吸虫尾蚴、虫卵和成虫的期别差异表达基因文库 . 经消减效率分析和三种文库克隆的 EST 的期别差异性鉴定,表明所建文库质量较高,为在整个基因组水平分离血吸虫的差异表达基因提供了重要材料 . 由三个文库选择 257 个插入片段大于 500 bp 的克隆测定了 EST 序列 . 同源性分析结果表明 257 个 EST 代表 182 种血吸虫基因,其中有 22 种为血吸虫已知基因,有 128 种为血吸虫已知 EST ,有 32 种为新发现的血吸虫基因 . 对 EST 编码蛋白的功能预测结果显示:尾蚴消减文库的基因多与运动、能量代谢、转录调节及致病性相关;虫卵消减文库的基因可能参与信号转导、细胞粘附、蛋白质和碳水化合物的代谢以及抗氧化反应;成虫消减文库的基因多参与蛋白质的合成、转运及分解代谢,参与虫体的运动等 . 大规模分离、分析血吸虫期别差异表达基因将对从分子水平去解读血吸虫的生长发育机制,筛选高效疫苗候选抗原、药物靶标及诊断制剂有重要意义 .

    Abstract:

    In order to understand the mechanism of growth and development of Schistosoma japonicum, the differentially expressed gene cDNA libraries of Schistosoma japonicum cercariae, egg and adult worm were constructed separately for the first time by SSH (suppressed subtractive hybridization) technique, which can be used to isolate and analyze differentially expressed genes on the whole genome scale. The results of subtractive efficiency and identification of stage difference among above three subtracted cDNA libraries indicated the quality were high. The 257 clones which inserted fragments more than 500 bp from above three cDNA libraries were sequenced. BLASTn results showed that these 257 ESTs represented 182 schistosome genes, including 22 schistosome known genes, 128 schistosome ESTs and 32 no match genes which may represent schistosome novel gene. Results of these ESTs function prediction indicated that differentially expressed genes from cercariae subtracted cDNA library were mainly involved in movement, energy metabolism, transcriptional regulation and pathogenesis; differentially expressed genes from egg were mainly involved in signal transduction, cell adhesion, protein and carbohydrate metabolism and response to oxidative stress; differentially expressed genes from adult worm were mainly involved in protein biosynthesis, transport and body movement. To isolate and analyze differentially expressed gene on large scale must provide the valuable information for understanding the mechanism of parasite growth and development on the molecular level, for screening highly effective antigen candidates, target of drug and diagnostic antigen.

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苑纯秀,冯新港,林矫矫,李 浩,陆 珂,石耀军,傅志强,刘金明,蔡幼民.日本血吸虫期别差异表达基因文库的构建及分析[J].生物化学与生物物理进展,2005,32(11):1038-1046

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