Molecular cloning and characterization of the UL31 gene from duck enteritis virus.
Using a combination of bioinformation analysis and Dot blot technique, a gene, designated hereafter as the duck enteritis virus (DEV) UL31 gene (GenBank accession number EF643559), was identified from the DEV CHv genomic library. Then, the UL31 gene was cloned and sequenced, which was composed of 933 nucleotides encoding 310 amino acids. Multiple sequence alignment suggested that the UL31 gene was highly conserved in Alphaherpesvirinae and similar to the other herpesviral UL31. Phylogenetic analysis showed that the gene had a close evolutionary relationship with the avian herperviruses, and DEV should be placed into a single cluster within the subfamily Alphaherpesvirinae. Antigen prediction indicated that several potential B-cell epitopes sites located in the UL31 protein. To further study, the UL31 gene was cloned into a pET prokaryotic expression vector and transformed into Escherichia coli BL21 (DE3). A 55 kDa fusion protein was induced by the further culture at 37 degrees C after addition of 0.8 mM IPTG. Polyclonal antibody raised against the recombinant UL31 from rabbit was prepared. A protein about 55 kDa that reacted with the antibody was detected in immunoblots of bacterial proteins, suggesting that the 55 kDa protein was the product of the UL31 gene. Immunofluorescence analysis revealed that the protein was localized in very fine punctate forms in the nuclei of infected cells. Our results may provide some insight for further research about the gene and also enrich the database of herpesvirus.
Xie
Wei
W
Avian Diseases Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Sichuan, China. xiew007@sohu.com
Cheng
Anchun
A
Wang
Mingshu
M
Chang
Hua
H
Zhu
Dekang
D
Luo
Qihui
Q
D018138:Alphaherpesvirinae / Q000145:classification / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D046090:Anseriformes / Q000821:virology*; D001483:Base Sequence; D015153:Blotting, Western; D002681:China; D003001:Cloning, Molecular; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D004926:Escherichia coli; D005455:Fluorescent Antibody Technique; D015698:Genomic Library; D015151:Immunoblotting; D008969:Molecular Sequence Data; D010802:Phylogeny; D017422:Sequence Analysis, DNA; D014764:Viral Proteins / Q000235:genetics*
D017931:DNA Primers; D014764:Viral Proteins