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意大利蜜蜂amPGI基因的克隆与表达分析
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引用本文:欧阳霞辉,徐文凯,郑相相,彭 帅,刘丽霞.意大利蜜蜂amPGI基因的克隆与表达分析[J].西北农业学报,2021,(12):1854~1862
DOI:10.7606/j.issn.1004-1389.2021.12.011
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作者单位
欧阳霞辉,徐文凯,郑相相,彭 帅,刘丽霞 (西北民族大学 生命科学与工程学院兰州 730030) 
基金项目:国家自然基金(31660628);甘肃省自然基金(1606RJZA058);中央高校基本科研业务费专项(31920210128)。
中文摘要:葡萄糖-6-磷酸异构酶(PGI)是糖酵解和几丁质合成途径中一种发挥重要作用的蛋白酶,为了探究意大利蜜蜂amPGI基因的分子特征及其在不同品级、不同发育时期的表达情况,采用RT-PCR技术克隆意大利蜜蜂葡萄糖-6-磷酸异构酶基因(amPGI),应用生物信息学方法分析其序列特征;进一步通过 RT-qPCR技术检测amPGI基因在意大利蜜蜂不同品级、不同发育时期的表达模式。结果显示,克隆获得amPGI基因序列全长1 674 bp,共编码557个氨基酸。序列分析发现,amPGI与大蜜蜂PGI氨基酸序列相似度达到 96.3%,蛋白结构以α-螺旋为主,且含有2个保守结构域和9个磷酸化位点,是一种结构稳定的非分泌型的亲水性胞内蛋白。表达模式显示amPGI基因在意大利蜜蜂不同品级、不同发育时期均有表达,且差异显著 (P<0.05),工蜂中幼虫期5日龄表达量最高,雄蜂中成虫期表达量最高,蜂王中幼虫期6日龄表达量最高,不同品级卵前期和蛹期表达量均较低,且卵的孵化和蛹的羽化过程中amPGI基因表达量都急剧增加。综上推测amPGI基因在意大利蜜蜂精巢和卵巢的发育及几丁质的合成过程中具有重要作用。
中文关键词:意大利蜜蜂  葡萄糖-6-磷酸异构酶  分子特征  表达模式
 
Cloning and Expression Analysis of Apis mellifera amPGI Gene
Abstract:Glucose-6-phosphate isomerase (PGI) plays an important role in the glycolytic pathway and Chitin Biosynthesis Pathwa. In order to explore the molecular characteristics and expression patterns of amPGI gene, the amPGI genes was cloned by RT-PCR , and its expression patterns of various grades at different developmental stages of Apis mellifera were analyzed. Moreover, its sequence characteristics were determined by use of bioinformatics methods.The results showed that the full length of the cloned sequence was 1 674 bp, containing a complete open reading frame which encoded 557 amino acids. The sequence of amPGI was highly homologous to that of A.dorsata,and the protein structure was dominated by α-helix. The amPGI protein had two conserved domains and nine phosphorylation sites, which was a non-secreted hydrophilic intracellular protein with stable structure. The results of RT-qPCR indicated that amPGI expression level was of significant differences at various grades and different developmental stages of A.dorsata(P<0.05). The workers had highest expression level in the 5 day-old larval,while the drones had highest expression level in the 6 day-old larvae and queens had it at adult stage. The amPGI gene expression was relatively low at the pre-egg stage and pupal stage of various grades, whereas, was sharply increased from the hatching of eggs and the pupal eclosion to the adult stage. The results suggest that the amPGI gene play a critical role in the development of testis and ovaries, and in chitin synthesis of A.mellifera.
keywords:Apis mellifera  Glucose-6-phosphate isomerase  Molecular characteristics  Expression pattern
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