| 家蚕滞育生物钟蛋白质EA4基因的cDNA克隆和序列分析 |
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| 引用本文： | 王玉军,徐世清,司马杨虎,吴阳春,刘莹,柳学广,丛海峰.家蚕滞育生物钟蛋白质EA4基因的cDNA克隆和序列分析[J].蚕业科学,2007,33(1):36-42. |
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| 作者姓名： | 王玉军 徐世清 司马杨虎 吴阳春 刘莹 柳学广 丛海峰 |
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| 作者单位： | 苏州大学生命科学学院,苏州,215123;苏州大学生命科学学院,苏州,215123;苏州大学生命科学学院,苏州,215123;苏州大学生命科学学院,苏州,215123;苏州大学生命科学学院,苏州,215123;苏州大学生命科学学院,苏州,215123;苏州大学生命科学学院,苏州,215123 |
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| 基金项目： | 国家自然科学基金
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国家重点基础研究发展计划(973计划) |
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| 摘 要： | 家蚕滞育生物钟是依赖卵内酯酶A4(EA4)的倒计时型生物钟。EA4是具时间间隔测定酶(time interval measuring enzyme,TIME)功能的蛋白质。根据家蚕EA4测定的氨基酸序列,以第3天龄雌蛹脂肪体和卵巢总RNA为模板,利用简并引物RT-PCR方法,得到308 bp的部分ea4序列,其1-144 nt和145-308 nt分别与家蚕全基因组鸟枪序列(whole genome shotgun sequence,WGS)AADK01019126.1的5595-5738 nt及WGS AADK01014467.1的8636-8473 nt有95%和98%的一致性。根据获得的ea4基因片段序列,利用生物信息学方法和家蚕基因组数据预测了ea4基因的完整cDNA序列,设计特异引物,以雌蛹脂肪体和卵巢RNA为模板,进行RT-PCR得到完整的cDNA。ea4基因的cDNA全长519 bp,其中1-48 bp为信号肽序列,49-519 bp编码的氨基酸与蛋白质纯化后测序EA4部分的一致性为98%,C末端存在3个氨基酸的差异,其中第156位为新确定氨基酸。
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| 关 键 词： | 家蚕 滞育 时间间隔测定酶 酯酶A4基因 cDNA序列 |
| 文章编号： | 0257-4799(2007)01-0036-07 |
| 修稿时间： | 2006-10-08 |
Cloning and Sequence Analysis of cDNA of Diapause Biological Clock Protein Esterase A4 of Bombyx mori |
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| WANG Yu-Jun,XU Shi-Qing,SIMA Yang-Hu,WU Yang-Chun,LIU Ying,LIU Xue-Guang,CONG Hai-Feng.Cloning and Sequence Analysis of cDNA of Diapause Biological Clock Protein Esterase A4 of Bombyx mori[J].Acta Sericologica Sinica,2007,33(1):36-42. |
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| Authors: | WANG Yu-Jun XU Shi-Qing SIMA Yang-Hu WU Yang-Chun LIU Ying LIU Xue-Guang CONG Hai-Feng |
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| Institution: | College of Life Science, Soochow University, Suzhou Jiangsu 215123, LIU Ying China |
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| Abstract: | The diapause biological clock protein of Bombyx silkworm is a count down clock which depends on esterase A4(EA4) in their eggs.EA4 was the first confirmed time interval measuring enzyme(TIME).A 308 bp of ea4 cDNA sequence was generated by RT-PCR with degenerate primers designed according to the determined amino acids and using the total RNA of fat body and ovary from 3-day old female pupa as the template.The consistence of 1-144 nt and 145-308 nt of the sequence with 5 595-5 738 nt of whole genome shotgun sequence(WGS)(AADK01019126.1) and 8 636-8 473 nt of WGS(AADK01014467.1) were 95% and 98%,respectively.According to the sequence of the ea4 gene fragment obtained,ea4 gene integrity cDNA sequence was forecasted by method of bioinformatics and the genome data of Bombyx mori.Then,a pair of special primers was designed according to the forecasted cDNA sequence of ea4,and the complete cDNA sequence was obtained by RT-PCR using the total RNA of fat body and ovary of female pupa as the template.The cDNA of ea4 gene was 519 bp in length,in which 1-48 bp was the signal peptide sequence.And the amino acid sequence encoded by 49-519 bp showed 98% similarity with that of EA4 protein,with 3 amino acids difference at the C-terminal,in which the 156th was the first defined one. |
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| Keywords: | Bombyx mori Diapause Time interval measuring enzyme ea4 gene cDNA sequence |
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