Abstract: DDX21 (DExD-box helicase 21) is a RNA helicase,which belongs to the DEAD-box family of RNA helicases.Previous reports showed DDX21 not only take part in the generation and processing of RNA,but also have functional roles in virus's replication.This experiment was conducted to obtain sequence of swine DEAD-box helicase 21 gene,and to study its prokaryotic expression.According to the predicted Sus scrofa DDX21 gene mRNA sequence in GenBank(XM_005657387.2),proper primers were designed.Total RNA was then extracted from porcine kidney passage cells(PK-15),and CDs sequence of the gene was amplified by RT-PCR using the primers.The sequence analysis results showed that the sequence of porcine DDX21 CDs was in length of 2 355 bp which encoded 784 amino acids (GenBank Accession No.KX396051); Compared with Bos taurus,Danio rerio,Homo sapiens,Mus musculus,Rattus norgicus and Xenopus tropicalis,homology of porcine DDX21 was 91.7%,57.5%,88.0%,82.3%,83.8%,and 48.9% at the amino acid level,respectively; Structural analysis with the SMART program indicated that porcine DDX21 contained a putative C-terminal GUCT domain.Similar GUCT domain domains had been identified in cattle,human,mouse and rat DDX21; Phylogenetic tree analysis showed that porcine DDX21 had the closest relationship with cattle DDX21.Additionally,prokaryotic expression vector pET28a-DDX21 was constructed for further study.After sequencing,the recombinant was transformed into Escherichia coli BL21 (DE3) competent cells.The SDS-PAGE experiment demonstrated that the recombinant objective protein appeared a molecular mass of approximately 90 kDa which was consistent with the anticipated size.When IPTG concentration was kept constant,5 h induction was optimal.Cloning of porcine DDX21 gene and expression in E.coli laid a foundation for the subsequent structural analysis and function research of this gene.

Key words: Porcine, DDX21 gene, Cloning, Sequence analysis, Prokaryotic expression

CLC Number: 

• Q78

• S828