大豆食心虫储存蛋白hexamerin基因的克隆与表达分析
Molecular cloning and expression analysis of the SpbHex(hexamerin gene) in soybean pod borer (Leguminivoraglycinivorella Matsumurav)
臧振原** 贾宝辉 李涵哲 李 娜 孟凡立 李文滨***
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作者单位：东北农业大学农学院 大豆生物学教育部重点实验室 哈尔滨 150030
中文关键词：大豆食心虫，hexamerin基因，基因克隆，表达分析
英文关键词：soybean pod borer, hexamerin gene, gene clone, expression analysis
中文摘要： 【目的】 储存蛋白是昆虫发育、变态和生殖过程中氨基酸的主要来源， Hexamerin是储存蛋白家族重要成员，在昆虫的生长发育中起着重要作用。为了研究hexamerin基因（SpbHex）在大豆食心虫Leguminivora glycinivorella Matsumurav生长发育过程中的作用，对大豆食心虫SpbHex基因进行克隆与表达分析。【方法】 利用RT-PCR和RACE技术克隆SpbHex的cDNA全长序列，并通过qPCR研究其在不同发育阶段和幼虫不同组织的表达情况。【结果】 SpbHex基因cDNA序列全长2 161 bp，其中开放阅读框2 112 bp，编码703个氨基酸。蛋白预测分子量84.15 ku。hexamerin基因在大豆食心虫不同发育阶段和不同组织中均有表达。在不同生长发育阶段中4龄幼虫的表达量较高，1龄和成虫的表达量较低；在不同组织中脂肪体的表达量较高，表皮中的表达量最低。【结论】 本研究克隆了大豆食心虫储存蛋白hexamerin 基因，并对其在大豆食心虫中表达模式进行分析，为进一步明确hexamerin基因在大豆食心虫生长发育中的作用奠定基础。
英文摘要： [Objectives] In insects, Storage Superfamily Proteins (SSPs) accumulate apparently to serve as sources of amino acids during metamorphosis, reproduction and development. Hexamerins are important SSPs. In order to study the function of the hexamerin gene in the development of soybean pod borer, the gene was cloned and its expression analyzed. [Methods] The full length of the cDNA sequence of the soybean pod borer hexamerin gene (SpbHex) was amplified using the reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). The expression of SpbHex in all developmental stages and tissues was analyzed using Realtime quantitative PCR. [Results] A soybean pod borer gene encoding hexamerin (SpbHex) was cloned. The full length of SpbHex was 2 161 bp with an open reading frame (OFR) of 2 112 bp. The ORF of SpbHex encodes a protein of 703 amino acids with a predicted molecular mass of approximately 84.15 ku. RT-PCR analyses indicates that SpbHex was expressed in all development stages, with the highest expression in fourth instar larvae and the lowest in first instar larvae and adults. Meanwhile, RT-PCR analyses indicates high quantities of SpbHex transcript in the fat body and low amounts in the cuticle. [Conclusions] The hexamerin gene of the soybean pod borer was cloned and its transcription profiles determined in all developmental stages and tissues. The present findings lays the foundation for further study of the function of SpbHex in the development of the soybean pod borer.