Molecular cloning and sequence analysis of fusion gene of paramyxovirus Tianjin strain
Objective To explore the origin and phylogenetic location of paramyxovirus Tianjin strain base on fusion(F)gene.Methods The genomic RNA of Tianjin strain was extracted and used as template.Ac- cording to GenBank Sendai viruses(SeV)sequences,two pairs of primers were designed and two fragments cover- ing the coding region of F gene were amplified by reverse transcription polymerase chain reaction(RT-PCR).The purified products of RT-PCR were cloned into pGM-T vector.Positive clones were identified by PCR and EcoRⅠenzyme digestion and then sequenced.The obtained nucleotide sequence and deduced amino acid sequence were compared with those of 15 SeV from GenBank,and phylogenetic trees based on the nucleotide sequences and amino acid sequences of F gene were constructed with Clustal X and TreeView software.Resulits The sequencing results demonstrated that F gene of Tianjin strain comprises 1698 nucleotides and encodes a protein of 565 amino acids, which is consistent with that of the SeV.neither insertion nor deletion was detected in the coding region.Homology analysis showed that Tianjin strain had a higher similarity to SeV BB1 strain.The identities of the F gene and the F protein between them were 94.5%,95.4%,respectively.The sequence comparison between Tianjin strain and other 14 SeV showed that the homology of uucleotide sequences was 85.3%-90.5%,and the identity of the de- duced amino acid sequences was 90.4%-94.5%.There are 13 unique amino acid substitutions in the F protein of Tianjin strain.The phylogenetic analysis based on the F gene revealed that 8 SeVs were divided into three different evolution branches.Tianjin strain becomes a unique branch,which is closely akin to the branch represented by BB1 strain and relatively farther akin to the branches represented by Z strain and Ohita strain.Condusion Pa- ramyxovirus Tianjin strain does not belong to any of the three known evolution branches of SeV but the fourth one.