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  

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

作　　者：于奎峰[1] 李红亮[1] 杨璞[2] 崔旭红[1] 祝增荣[2] 商晗武[1] 

机构地区：[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

出　　处：《中国水稻科学》2010年第6期575-580,共6页Chinese Journal of Rice Science

基　　金：国家自然科学基金资助项目(30571241);国家973计划资助项目(2009CB119200);浙江省重大农业科研专项资助项目(2006C12020)

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1727bp和1730bp,均包含1344bp的完整开放阅读框(ORF),编码447个氨基酸,理论分子量约50kD,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(3.14×107copies/μL±0.66×107copies/μL)显著高于两性生殖型稻水象甲(5.08×106copies/μL±0.47×106copies/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。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 1727bp for the parthenogenetic RWW,and 1730bp for the sexual RWW.Both cDNA sequences contained a 1344bp open reading frame encoding 447amino acid residues.The putative molecular weight of these amino acid residues was approximately 50kD at an isoelectric point of 4.54.The clonedβ1-tubulin amino acids shared a high similarity（95% to 100%）with theβ-tubulin homologues from other insects.Absolute quantification revealed that theβ1-tubulin was expressed at a significantly higher level in the ovary of the parthenogenetic RWW（3.14×107 copies/μL±0.66×107copies/μL）than the sexual RWW（5.08×106copies/μL±0.47×106copies/μL）.It is suggested that the differential expression ofβ1-tubulin in the parthenogenetic RWW may play an important role in the formation of the mitotic spindle.

关 键 词：稻水象甲 β1-微管蛋白 CDNA克隆 定量表达 

分 类 号：S435.112[农业科学—农业昆虫与害虫防治]