Molecular Cloning and Expression of the Hemagglutinin ( H ) Gene of the Canine Distemper Virus

• Main Authors: Ta-Chun Chen, 陳大鈞
• Other Authors: Ming-Huei Liao
• Format: Others
• Language: zh-TW
• Published: 2004
• Online Access: http://ndltd.ncl.edu.tw/handle/30271881483247550965

碩士 === 國立屏東科技大學 === 獸醫學系 === 92 === Canine distemper (CD) appears since 1760 in Europe, but canine distemper virus (CDV) was identified in 1906. CDV is a highly contagious viral pathogen, which is a very important viral pathogen of dogs. It often infect 3~6 month old puppies, but all ages dogs of could also be infected causing acute to subacute systemic diseases with high mortality rates develop. Targets of infection for CDV are mainly mucous membranes and lymphoid tissues throughout the body; thus, the disease is typically characterized by manifestations of pyrexia, anorexia, nasal mucopurulent discharge, conjunctivitis, and diarrhea. Skin pustules, hyperkeratosis, and central nervous system signs are also seen in some dogs. Several papers pointed out that nucleotide sequence analysis of actual field viruses had undergone significant genetic changes relatived to currently used vaccine strains, and CDV strains genetically different from vaccine strains may have caused the disease in vaccinated dogs, and the highest antigenic variation has been found in the H gene. Therefore, the goal of this study is to clone and express the partial H gene of CDV from the CD vaccine strain, and to identify the variation of H gene of vaccine strains and wild type strains of CDV in Taiwan. The partial H gene was amplified by reverse transcription-polymerase chain reaction (RT-PCR) with specific primer, then the sequenced and cloned into pET32a expression vector. Purified crude protein was checked by SDS-PAGE, immunodotting test and Western blotting assay by polyclonal antibodies produced in rabbit. In the result, dot blotting assay idicated that the antibodies could specific bind with the protein that we expressed and with protein from CDV vaccine.