[Cloning and expression of novel swine gene BCL-G(L) in E.coli and preparation of its polyclonal antibody in guinea pigs].
OBJECTIVE
In order to express a novel gene named as BCL-G(L); of swine in E.coli and prepare its polyclonal antibody.


METHODS
The contig sequence of the gene was predicted and in silicon cloned by blasting the human BCL-G(L); in swine ESTs database in NCBI. The cloning sequence was obtained by RT-PCR from swine spleen. The cloning sequence was identified by sequencing and compared with the contig sequence. Then the gene was cloned into a prokaryotic expression vector pET-32a to construct a recombinant plasmid named as pET32a-BCL-G(L);. The fusion protein pET32a-BCL-G(L); was expressed in E.coli BL21 and purified using a His-tag fusion protein purification kit. Then guinea pigs were immunized with the purified protein to get the specific polyclonal antibody.


RESULTS
The titer of the antibody was 1:800 detected by ELISA. The protein BCL-G(L); can be specifically detected by western blot assay using the polyclonal antibody.


CONCLUSIONS
The novel swine gene BCL-G(L); was cloned and expressed in E.coli and its polyclonal antibody was prepared successfully.
Research Laboratory of Virology, College of Veterinary Medicine, Northwest Agriculture & Forestry University, Yangling 712100, China. pengfei519@126.com
Peng-Fei
PF
Jiang
Jin-Suo
JS
Cao
Chao
C
Liu
Hai-Ping
HP
Zhao
De-Li
DL
Zhang
D000818:Animals; D000906:Antibodies / Q000276:immunology*; D000918:Antibody Specificity; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D005822:Genetic Vectors / Q000235:genetics; D006168:Guinea Pigs; D006801:Humans; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D015183:Restriction Mapping; D013552:Swine / Q000235:genetics*
D000906:Antibodies; D018076:DNA, Complementary; D019253:Proto-Oncogene Proteins c-bcl-2; D007544:Isopropyl Thiogalactoside