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摘要
beta-N-acetylglucosaminidase plays an important role in insect chitin degradation. In this study, a novel cDNA sequence encoding beta-N-acetylglucosaminidase (MbNAG) was cloned from cabbage moth, Mamestra brassicae L. The cDNA sequence, which was 2445 by in length, contained an open reading frame of 1782 by coding for a poly peptide of 594 amino acid residues. The MbNAG open reading frame was cloned into the pET-21b prokaryotic expression vector, and then over-expressed in Escherichia coli BL21 (DE3) cells. After purification and renaturation, the beta-N-acetylglucosaminidase enzymatic activity was detected. Transcript analyses on MbNAG sequence during different developmental stages and in specific tissues were carried out by RT-qPCR. The MbNAG mRNA level was significantly higher of the second day in the prepupal stage than at any other time and occurred mainly in the midguts and salivary glands compared with in other tissues, including fat bodies, integuments, malpighian tubules, foreguts and hindguts during the feeding stage of the 5th instar larvae. The highest expression level of MbNAG mRNA occurred 48 h after an in vivo injection of 20-hydroxyecdysone. RNA interference reduced the MbNAG mRNA expression level 60% at 48 h and 67% at 72 h, respectively, and induced both abnormal molting phenotypes and a high mortality rate in M. brassicae larvae. These results suggest that MbNAG plays an essential role in the molting process of M. brassicae.
- 出版日期2016-9
- 单位东北农业大学
