Isolation of infectious bursal disease virus in cell culture, detection and characterization by RT-PCRSSCP analysis
Gaba, Amit; Makadiya, N.R.; Joshi, C.G. and Jhala, M.K. (2007) Isolation of infectious bursal disease virus in cell culture, detection and characterization by RT-PCRSSCP analysis. Indian Journal of Poultry Science, 3 (42). pp. 339-342.
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The study was aimed at isolation of infectious bursal disease virus (IBDV), detection by RT-PCR and genetic characterization by single strand conformational polymorphism (SSCP). BGM-70 cwell monolayers inoculated with two field bursal samples resulted in isolation of the virus after five succesive passages. Distinct CPE charcaterized by rounding and clumping of cells, extensive cytolasmic granularity and detachmenet of the cells was obsorved, which was similar to that produced by vaccine isolated during first passage. RT-PCR performed for viral RNA extracted by Trizol reagent from cell culture supernatant and infected monolayers, using P1 and P2 pair of primers specific to hypervariable region of VP2 gene, resulted in generation of a targeted amplicon of 643 bp, confirming the growth of IBDV in cell culture. On SSCP analysis, RT-PCR amplified products of both the field isoalted and vaccine virus revealed similar migration pattern, suggesteing absence of any nucleotide sequence variation among them with respect to the targated gene segment.
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