关键词: 亚洲玉米螟, 酚氧化酶原激活蛋白酶, 基因克隆, 序列分析, 亲缘关系

Abstract:

In order to explore the molecular mechanism of expression and regulation of prophenoloxidase activating proteinase (PAP) in larvae of Ostrinia furnacalis (Guenée), the sequences of conserved regions of PAP genes in different insects were used to design the degenerate primers, and a cDNA fragment with the size of 509 bp encoding 169 amino acids was amplified from the 5th larvae of O. furnacalis by RT-PCR. The fragment has a predicted molecular weight of 18.7 kD and pI value of 5.1. A conserved catalytic triad (H, D, S) in the serine protease like domain was found, but the clip domain did not exist. BlastP analysis showed that the amino acid sequence of the cloned fragment from O. furnacalis had the highest identity (47%) to those of Manduca sexta PAP-3 and Anopheles gambiae Clip B1 protein, and 45%, 45%, 44%, 43% and 41% identity to those of M. sexta PAP-2, Bombyx mori PPAE, Spodoptera litura PPAE-3, Samia cynthia ricini PAP and Drosophila melanogaster serine protease-7, respectively. Phylogenetic analysis showed that the genetic relationship of O. furnacalis, M. sexta PAP-3 and S. litura PPAE-3 was closer, while that of O. furnacalis, Drosophila melanogaster serine protease-7 and Anopheles gambiae Clip B1 protein was more distant. The results suggest that the cloned cDNA fragment of O. furnacalis larvae is a part of the PAP gene nucleotide sequence at the C-terminus.

Key words: Ostrinia furnacalis, prophenoloxidase activating proteinase, gene cloning, sequence analysis, genetic relationship