Determination of the complete nucleotide sequence of Clone IR 12 strain derived from the LaSota strain of Newcastle Virus of Razi Institute

Document Type : Full Research Paper

Authors

Razi Serum and Vaccine Research Institute, Karaj, Iran

Abstract

Newcastle disease (ND) is an important viral disease of poultry causing severe economic losses and drop in egg production in broiler and layers flocks. Despite vaccination with both live attenuated and inactivated NDV strains to immunize birds, ND is still a serious problem in poultry industry especially in developing countries like Iran. plaque purified NDV strain clone IR12 produced by Razi Institute was propagated in 10-day-old SPF emberyonated chicken eggs, then allantoic fluid was harvested. The virus was purified using sucrose-gradient. RNA was extracted and cDNA spanning the whole genome was synthesized using two GSP primers. PCR was run using 7 sets of primers covering the whole cDNA.The PCR products were all gel extracted and ligated to pJET1.2 vector by 3-1 ratios. Successful cloning of each insert was confirmed using restriction enzyme BglII that digests areas flanking the insert in pJET. After confirmation, the inserts were sequenced using Sanger sequencing with pJEt specific primers as well as primers designed to bind to middle regions of each fragment. Sequence of the whole 15186 bp genome was determined by assembling sequences using MEGA 5software. Location of each start (S) and ending (E) gene signals as well as all ORF start and termination codons were determined.

Keywords

References

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History

  • Receive Date: 14 October 2019
  • Revise Date: 07 September 2020
  • Accept Date: 15 September 2020

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