Acta Agriculturae Boreali-Sinica ›› 2022, Vol. 37 ›› Issue (1): 195-201. doi: 10.7668/hbnxb.20192510

Special Issue: Biotechnology

• Resources & Environment·Plant Protection • Previous Articles     Next Articles

Cloning and Prokaryotic Expression of a clip-type Serine Protease Gene from Mythimna separate

LIAN Kaiqi1, JI Shuang1,2, ZHOU Lingling1, SHI Zhiqi1, WANG Fei1,2, ZHANG Yuanchen1,2   

  1. 1. School of Biotechnology and Food Science,Anyang Institute of Technology,Anyang 455000,China
    2. Taihang Mountain Forest Pests Observation and Research Station of Henan Province,Linzhou 456550,China
  • Received:2021-07-19 Published:2022-02-28


To clone the full gene sequence of Mythimna separate serine protease,and express it in vitro,Mythimna separata was used as the research object,the partial gene of serine protease was amplified by RT-PCR,and further the whole gene was obtained by RACE technology.The obtained gene sequence and deduced protein sequence of serine protease were analyzed using Blast software and other softwares.The gene of serine protease was expressed using the Escherichia coli expression system.The results showed that the full serine protease gene from Mythimna separate was successfully cloned and named as MsPG,with the sequence length of 2 010 bp,and the open reading frame of 1 593 bp,encoding 530 amino acids.Its protein had the molecular weight of 67.6 ku,and the isoelectrical point of 7.63,and owned a clip-type domain containing 6 cysteine,which was speculated as a clip-type serine protease.The amino acid sequence of MsPG had identities of 70.00%—91.92% to those of other Lepidoptera insects,and showed the highest identity(91.92%)to that of Heliothis virescens(GenBank No.PCG78401.1).The recombinant plasmid pET30a(+)-MsPG was successfully constructed and expressed by 0.1 mmol/L IPTG induction in E.coli prokaryotic expression system,with the highest expression at 25 ℃.In conclusion,MsPG has conserved functional sites in the serine protease family and can be prokaryotically expressed in vitro.

Key words: Mythimna separate, Serine protease, Gene cloning, Sequence analysis, Prokaryotic expression

Cite this article

LIAN Kaiqi, JI Shuang, ZHOU Lingling, SHI Zhiqi, WANG Fei, ZHANG Yuanchen. Cloning and Prokaryotic Expression of a clip-type Serine Protease Gene from Mythimna separate[J]. Acta Agriculturae Boreali-Sinica, 2022, 37(1): 195-201. doi: 10.7668/hbnxb.20192510.

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