In Vitro Expression of Filarial SXPI Gene for the Development of a Nucleic Acid Based Vaccine
The objectives of this study were to clone gene that encode filarial SXPI protein followed by in vitm expression of the protein. The Special Programme for Research and Training in Tropical Diseases (TDR) WHO has advocated SXPl as one of the vaccine candidate to curb filarial infection. SXPl antig...
Saved in:
Main Author: | |
---|---|
Format: | Thesis |
Language: | English |
Published: |
2005
|
Subjects: | |
Online Access: | http://psasir.upm.edu.my/id/eprint/6316/1/FPSK%28M%29_2005_2.pdf |
Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
Summary: | The objectives of this study were to clone gene that encode filarial SXPI
protein followed by in vitm expression of the protein. The Special Programme
for Research and Training in Tropical Diseases (TDR) WHO has advocated
SXPl as one of the vaccine candidate to curb filarial infection. SXPl antigen
has been reported to confer protective immunity, causing reduction of
microfilaraemia levels in jirds (Meriones unguiculatus) blocking subsequent
Brugia malayi infection. In this study, the gene that encode SXPl antigen was
517 bp in length and was extracted and amplified from the infective stage (L3)
of subperiodic Brugia malayi. The gene was successfully cloned into
replication vector p ~ ~ ~ 2(Inv. it1rog en) followed by subcloning into
mammalian expression vector pVAXl (Invitrogen). The presence of SXPl
gene in - both vectors were validated by polymerase chain reaction (PCR),
restriction enzymes analysis (RE) and finally by automated sequencing. The
cloned SXPl in pVAX was designated as pVAXISXP1. The plasmid bearing
SXPI gene was transfected into two types of animal cell lines (COS-7 and
CHO) using Polyfect Transfection Reagent (Qiagen). The successful
expression of targeted gene in the mammalian cell lines were determined by
RT-PCR and Western Blotting. The PCR product of the transfected cells was
517 bp on the agarose gel. In addition, the -20 kDa of expressed SXPl
protein was detected on nitrocellulose membrane by rabbit polyclonal
antibody against the SXPI protein. This study has successfully established
the ground work for future deliberations towards the development of antibrugia
transmission blocking genetic vaccine. |
---|