论文部分内容阅读
Objective:To discuss the possibility of expressing the haemagglutinin-neuraminidase (HN) protein in prokaryotic system such asEscherichia coli(E. coli) cells by cloning the full length HN gene.Methods:The full lengthHN gene of Newcastle Disease Virus(NDV) of size1 734bp was preciously isolated byRT-PCR. The sequence was assessed and submitted to Nucleic Acid Databank(NCBI) and the gene ID wasEU215390.1 after cloning and sequencing. Now the assessed HN gene was subcloned into pET 32 a+ expression vector for production theHN protein inE. coli, BL21 (DE3) PLYSS cells following standard protocols. The crude lysate protein from the induced positive clone was size assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and their haemagglutination(HA) property against chickenRBC was assessed by standard microHA test.Results: The molecular size of the fullHN gene ofNDV as assessed by cloning and digesting the positive clone to release the insert was 1.7kb. The expressed protein in both crude and pure form was assessed to be63 kDa and 81 kDa, respectively. TheHA activity of the crude protein of the positive clone was1 in40.Conclusions:This finding indicates that the fusion protein retains the biological activity of native protein in the crude form and therefore could be used as a diagnostic reagent for antibody detection and for routine assessment of immune status in commercial layer forms.