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OJVRTM
Online Journal of Veterinary
Research©
Volume 17 (5): 261-269, 2013
Partial
cloning
and expression of HA1 and HA2 gene of
avian
influenza H9N2 gene in Lactococcus
lactis.
Karimi Madab
M (DVM)1, Dadras H
(DVM, PhD)2,
Hosseini A(DVM, PhD)3,
Hosseinian
SA(DVM)4, Sepehrimanesh
M (DVM).5
1,2,4 Department(s)of
Avian Medicine, 3Immunology, 5Biochemistry and
Basic Sciences,
School of Veterinary Medicine, Shiraz University, Iran.
ABSTRACT
Karimi Madab M, Dadras
H, Hosseini A, Hosseinian
SA, Sepehrimanesh M., Partial
cloning and expression
of HA1 and HA2 gene of avian influenza H9N2 in Lactococcus
lactis. Onl
J Vet Res., 17 (5): 261-269, 2013. HA1 and HA2 gene antigenic sites of virus were
amplified and
cloned for expression in Lactoccocus lactis
for
possible use in a live vaccine. The target gene was amplified
by RT-PCR using
two pairs of primers and cloned into pNZ8148 and pNZ8110 L. lactis
expression vectors for cytoplasmic
and secretory heterologous protein to yield
pNZ8148-HA/C and pNZ8110-HA/S,
respectively. After preliminary transformation into E.
coli MC1061 the
plasmids were extracted and transformed to L. lactis
NZ9000 by electroporation.
During each step the
location and accuracy of genes sequence were confirmed by PCR,
restriction
enzyme analysis and sequencing. SDS-PAGE and silver staining
of purified
His-tagged proteins from recombinant bacteria revealed that
the HA was
successfully expressed in cytoplasmic
and secretory forms.
Key
words: Cloning, expression, Lactococcus
lactis NZ9000,
pNZ8110, pNZ8148, H9N2