西伯利亚蝗抗菌肽基因的克隆及表达特性研究

Cloning and expression of the attacin gene in Gomphocerus sibiricus

【目的】扩增西伯利亚蝗Gomphocerus sibiricus抗菌肽(attacin)基因,并对其进行生物信息学分析,同时分析西伯利亚蝗不同组织中抗菌肽基因的差异表达。【方法】基于RACE技术从西伯利亚蝗中克隆得到抗菌肽,利用生物信息学软件对西伯利亚蝗抗菌肽的一级结构、二级结构、三级结构、系统进化和亚细胞定位进行分析。采用实时荧光定量PCR技术检测了西伯利亚蝗抗菌肽基因在马氏管、中肠、后肠、脂肪体和唾液腺中的相对表达量。【结果】克隆获得的抗菌肽基因的开放阅读框为282bp,编码93个氨基酸,其编码蛋白预测理论分子量为9.98ku,等电点为9.26。抗菌肽氨基酸序列中存在信号肽序列。基因定量分析显示,抗菌肽基因在西伯利亚蝗的中肠中表达量最高。【结论】attacin在西伯利亚蝗成虫不同组织中差异表达,提示其消化道在防御病原微生物的入侵中起着重要的作用。

[Objectives]The attacin gene of Gomphocerus sibiricus was amplified and sequenced and its differential expression in different tissues was quantified.[Methods]The attacin gene was cloned using RACE technology and its primary structure,secondary structure,tertiary structure,phylogeny and subcellular localization,analyzed with bioinformatics software.Real-time quantitative PCR(RT-qPCR)was used to detect the relative expression of the gene in the Malpighian tubules,midgut,hindgut,fat body and salivary glands.[Results]The cloned attacin gene contained an open reading frame of 282 bp and encoded 93 amino acid residues.Its predicted molecular weights is 9.98 ku,and its isoelectric point is 9.26.Sequencing revealed signal peptide sequences.The gene was most highly expressed in the midgut.[Conclusion]Differences in expression levels of the attacin gene in different adult tissues of G.sibiricus suggests that the digestive tract plays an important role in preventing the invasion of pathogenic microorganisms.