[Expression of GST-PRL-2 fusion protein in prokaryotic cells and preparation of Hen egg yolk immunoglobulin (IgY) against PRL-2].
OBJECTIVE
To express phosphatase 2 of regenerating liver (PRL-2) fusion protein in E.coli and to prepare specific hen egg yolk immunoglobulin(IgY) against PRL-2.


METHODS
A full-length human PRL-2 gene coding sequence was cloned into expression vectors pGEX-4T-2 and pET21a, then transformed into E.coli. Fusion protein GST-PRL-2 was expressed in E.coli via IPTG induction. The expressed proteins were purified from lysates through Glutathione Sepharose 4B and the Ni-NTA agarose columns, respectively. Egg-laying hens were immunized using the purified GST-PRL-2 with Freund's complete or incomplete adjuvant. The specificity of the resulting antibody was identified by Western blot.


RESULTS
A high level of expression of target protein was detected by Western blot after IPTG induction and purified protein was obtained through Glutathione Sepharose 4B and the Ni-NTA affinity chromatography agarose columns, respectively. Western blot analysis showed that the anti-PRL-2 polyclonal antibody can recognize 6 x His-PRL-2 fusion protein.


CONCLUSIONS
The hen egg yolk immunoglobulin(IgY) against PRL-2 expressed in E.coli has good specificity which provides an useful reagent for the detection of PRL-2.
Guo
Ai-lin
AL
Department of Pathology, Zhongshan Medical College, SUN Yat-sen University, Guangzhou 510080, China. alinguo@yahoo.com.cn
Cao
Yun-xin
YX
Wang
Wen
W
Ding
Bo
B
Huang
Yu-qiang
YQ
Wen
Jian-ming
JM
D000818:Animals; D000918:Antibody Specificity; D002645:Chickens / Q000276:immunology*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D005982:Glutathione Transferase / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D007114:Immunization; D007136:Immunoglobulins / Q000276:immunology / Q000302:isolation & purification*; D010063:Ovum / Q000276:immunology; D011387:Prokaryotic Cells / Q000378:metabolism; D054648:Protein Phosphatase 2; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D014170:Transformation, Genetic
C016512:IgY; D007136:Immunoglobulins; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; D054648:Protein Phosphatase 2; C487561:PTP4A2 protein, human; D017027:Protein Tyrosine Phosphatases