Molecular Cloning and Quantitative Expression of a FullLength cDNA Encoding β1Tubulin in the Ovary of Rice Water Weevil, Lissorhoptrus oryzophilus (Coleoptera: Curculionidae)

doi:10.3969/j.issn.1001-7216.2010.06.003

Chinese Journal of Rice Science

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Molecular Cloning and Quantitative Expression of a FullLength cDNA Encoding β1Tubulin in the Ovary of Rice Water Weevil, Lissorhoptrus oryzophilus (Coleoptera: Curculionidae)

YU Kui-feng¹; LI Hong-liang¹; YANG Pu²; CUI Xu-hong¹; ZHU Zeng-rong²;SHANG Han-wu^1，*

¹College of Life Sciences, China Jiliang University/Zhejiang Provincial Key Laboratory of Biometrology and Inspection & Quarantine, Hangzhou 310018, China； ²Key Laboratory of Molecular Biology of Crop Pathogens and Insects, Ministry of Agriculture/Institute of Insect Sciences, Zhejiang University, Hangzhou 310029, China； *Corresponding author, E-mail: hwshang@cjlu.edu.cn

Received:1900-01-01 Revised:1900-01-01 Online:2010-11-10 Published:2010-11-10

稻水象甲卵巢内β1微管蛋白基因cDNA全长的克隆及定量表达分析

于奎峰¹; 李红亮¹; 杨璞²;崔旭红¹ ;祝增荣²; 商晗武^1，*

¹中国计量学院生命科学学院/浙江省生物计量及检验检疫技术重点实验室，浙江杭州 310018； ²浙江大学昆虫科学研究所/农业部作物病虫分子生物学重点开放实验室，浙江杭州 310029； *通讯联系人， E-mail: hwshang@cjlu.edu.cn

Abstract

Abstract:

A fulllength cDNA encoding β1tubulin from the parthenogenetic rice water weevil (RWW), Lissorhoptrus oryzophilus in China and from the sexual RWW in the United States was cloned by RACE and RTPCR, respectively. The full cDNA sequence was 1727 bp for the parthenogenetic RWW, and 1730 bp for the sexual RWW. Both cDNA sequences contained a 1344 bp open reading frame encoding 447 amino acid residues. The putative molecular weight of these amino acid residues was approximately 50 kD at an isoelectric point of 4.54. The cloned β1tubulin amino acids shared a high similarity (95% to 100%) with the βtubulin homologues from other insects. Absolute quantification revealed that the β1tubulin was expressed at a significantly higher level in the ovary of the parthenogenetic RWW (3.14×107 copies/μL±0.66×107 copies/μL) than the sexual RWW (5.08×106copies/μL±0.47×106 copies/μL). It is suggested that the differential expression of β1tubulin in the parthenogenetic RWW may play an important role in the formation of the mitotic spindle.

Key words: Lissorhoptrus oryzophilus, β1tubulin, cDNA cloning, quantitative expression

摘要： 利用cDNA末端快速扩增技术（RACE）和反转录聚合酶链式反应（RTPCR）分别对中国的孤雌生殖型和美国的两性生殖型稻水象甲卵巢内β1微管蛋白基因进行了克隆，获得的cDNA全长分别为1727 bp和1730 bp，均包含1344 bp的完整开放阅读框(ORF)，编码447个氨基酸，理论分子量约50 kD，等电点4.54。同源性分析表明，克隆的β1微管蛋白基因与其他昆虫的β微管蛋白基因在氨基酸水平上是高度同源的，相似性介于95%~100%。荧光定量PCR分析表明，β1微管蛋白在孤雌生殖型稻水象甲卵巢内的表达量（3.14×107 copies/μL±0.66×107 copies/μL）显著高于两性生殖型稻水象甲（508×106 copies/μL± 047×106 copies/μL）。推测该基因的差异表达对于稻水象甲孤雌生殖过程中纺锤体的组装具有重要意义。

关键词: 稻水象甲, β1微管蛋白, cDNA克隆, 定量表达

YU Kui-feng,LI Hong-liang,YANG Pu,CUI Xu-hong,ZHU Zeng-rong,SHANG Han-wu. Molecular Cloning and Quantitative Expression of a FullLength cDNA Encoding β1Tubulin in the Ovary of Rice Water Weevil, Lissorhoptrus oryzophilus (Coleoptera: Curculionidae)[J]. Chinese Journal of Rice Science, DOI: 10.3969/j.issn.1001-7216.2010.06.003 .

于奎峰, 李红亮, 杨璞,崔旭红 ,祝增荣, 商晗武,. 稻水象甲卵巢内β1微管蛋白基因cDNA全长的克隆及定量表达分析[J]. 中国水稻科学, DOI: 10.3969/j.issn.1001-7216.2010.06.003 .

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Molecular Cloning and Quantitative Expression of a FullLength cDNA Encoding β1Tubulin in the Ovary of Rice Water Weevil, Lissorhoptrus oryzophilus (Coleoptera: Curculionidae)

稻水象甲卵巢内β1微管蛋白基因cDNA全长的克隆及定量表达分析

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