[Expression of c-terminal of Parkin in E. coli and the preparation of antiserum].
OBJECTIVE
To clone and express 3'-terminal of Parkin gene in E. coli, and prepare its antiserum for further study.


METHODS
The glutathion-sulfate-transferase (GST) fusion expression plasmid of 3'-terminal of Parkin gene (937-1959 bp) was constructed and transferred to JM 105. After being treated with Triton-100 (1%) and Tween-20 (1%) and purified with affinity chromatograph, GST-Parkin C was used to immunize New Zealand rabbits to acquire antiserum. Antiserum was analysed with immunoblot.


RESULTS
The GST-Parkin C protein was expressed in JM 105, existing in the form of inclusion body with a molecular weight of around 42 kD; The purity of GST-Parkin C was up to 95%; the titer of antiserum was 1:64; Immunoblotting showed that the prepared antiserum could react specifically with 51.6 kD protein extracted from the mouse brain.


CONCLUSIONS
A high level of expression of GST-Parkin C is obtained in JM 105, and its antiserum can be prepared successfully.
Ou Yang
Jue
J
National Laboratory of Medical Genetics, Central South University, Changsha 410078, China.
Xia
Jun
J
Zheng
Duo
D
D000818:Animals; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D005982:Glutathione Transferase / Q000235:genetics; D007106:Immune Sera / Q000096:biosynthesis / Q000276:immunology; D008025:Ligases / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D051379:Mice; D010957:Plasmids / Q000235:genetics; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D044767:Ubiquitin-Protein Ligases
D007106:Immune Sera; D011993:Recombinant Fusion Proteins; D044767:Ubiquitin-Protein Ligases; C111567:parkin protein; D005982:Glutathione Transferase; D008025:Ligases