Record Details

Cloning and expression of fusion (F) and haemagglutinin-neuraminidase (HN) genes of Newcastle disease virus in insect host (Sf9cells)

Indian Agricultural Research Journals

View Archive Info
 
 
Field Value
 
Title Cloning and expression of fusion (F) and haemagglutinin-neuraminidase (HN) genes of Newcastle disease virus in insect host (Sf9cells)
 
Creator GOWRAKKAL, MAHESWARAPPA
VIJAYARANI, K
KUMANAN, K
 
Subject Newcastle disease virus
Specific pathogen free (SPF) embryonated chicken eggs
Recombinant antigens
 
Description The lentogenic strain of Newcastle disease virus (NDV) was propagated in 9–11 day-old specific pathogen free (SPF) embryonated chicken eggs. Allantoic fluid was collected after the death of embryo, the obtained allantoic fluid showed virus titre (HA) of 1:256. Viral RNA was extracted from allantoic fluid by Trizol method, complementary RNA was synthesized by reverse transcriptase enzyme (superscript-III) and full length recombinant fusion and recombinant haemagglutinin- neuraminidase gene was amplified by PCR using gene specific primers. Gel purified PCR products were cloned into pTriEx neo 1.1 expression vector and transformed into E. coli BL-21 DE3 cells. The presence of insert in recombinant colonies was confirmed by colony PCR, restriction digestion and sequencing of insert released products. Cloned recombinant plasmids of fusion and haemagglutinin neuraminidase genes were expressed in Sf9 cells using baculovirus expression vector system. After 72 h of transfection plaque assay was done to confirm the presence of recombinant virus and then positive recombinant viruses were transfected to insect cells. The expression of recombinant proteins in Sf9 cells was confirmed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS- PAGE), immunoblotting and immunofluorescent assays (IFA).
 
Publisher Indian Council of Agricultural Research
 
Date 2014-02-12
 
Type info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
 
Format application/pdf
 
Identifier https://epubs.icar.org.in/index.php/IJAnS/article/view/37835
10.56093/ijans.v84i2.37835
 
Source The Indian Journal of Animal Sciences; Vol. 84 No. 2 (2014); 162–165
2394-3327
0367-8318
 
Language eng
 
Relation https://epubs.icar.org.in/index.php/IJAnS/article/view/37835/16967
 
Rights Copyright (c) 2014 The Indian Journal of Animal Sciences
https://creativecommons.org/licenses/by-nc-sa/4.0