小地老虎β-N-乙酰葡萄糖胺糖苷酶基因cDNA序列的克隆及mRNA组织特异性表达

β-N-乙酰葡萄糖胺糖苷酶是昆虫几丁质代谢中一种关键酶,在昆虫的多种生理活动中发挥重要作用。本文以小地老虎Agrotis ipsilon Hufngel预蛹期幼虫为材料提取总RNA,利用RT-PCR和RACE技术,扩增得到其β-N-乙酰葡萄糖胺糖苷酶基因的cDNA序列。该序列含有2701个碱基,包括一个1788个碱基的开放阅读框,编码一个含595个氨基酸的蛋白,分子量约为68.3ku,等电点为5.48。推导得到的氨基酸序列与其他昆虫,尤其是鳞翅目昆虫的β-N-乙酰葡萄糖胺糖苷酶高度同源。基因cDNA序列已经登录GenBank并获得登录号GU985280。该基因在小地老虎取食期和预蛹期的马氏管、中肠、脂肪体、体壁和去中肠虫体中均含有mRNA水平的表达。

cDNA cloning and mRNA expression of β-N-acetylglucosaminidase in different tissues of Agrotis ipsilon Hufngel

β-N-acetylglucosaminidase is an enzyme that can degrade chitin in insects. Total RNA was isolated from the prepupal stage of Agrotis ipsilon Hufngel. The eDNA sequence was cloned by RT-PCR and rapid amplification of eDNA ends（RACE）. The eDNA,2 701 base pairs in length,contained an open reading frame of 1 788 base pairs coding for a polypeptide of 595 amino acid residues with a predicted molecular weight of 68.3 ku and pI 5.48. The predicted protein shared extensive similarities with those from other insects,expecially Lepidopteran insects. The eDNA sequence has been deposited in GenBank with the accession No. GU985280. RT-PCR results indicate that β-N-acetylglueosaminidase mRNA was expressed in the malpighian tubules, midgut, fat bodies and integument.