Expression of recombinant ribosome inactivating protein MAP30 in E.coli and its biological activity / 浙江大学学报·医学版
Journal of Zhejiang University. Medical sciences
;
(6): 264-271, 2010.
Article
in Chinese
| WPRIM
| ID: wpr-259205
ABSTRACT
<p><b>OBJECTIVE</b>To clone and produce ribosome inactivating protein MAP30 from the seeds of Momordica charantia L(bitter melon), and to evaluate the biological activity of the recombinant protein.</p><p><b>METHODS</b>The DNA sequence encoding MAP30 was cloned from the fresh seeds of Momordica charantia by PCR, the target DNA fragments were sequenced after T-A cloning. The expression plasmid was constructed by inserting the MAP30 fragment into vector pET30a. MAP30 was expressed in E.coli by addition of IPTG into final concentration of 1.0 mmol/L. The recombinant MAP30 was identified by SDS-PAGE, and the biological activity of MAP30 protein was evaluated by using MTT assay in cancer cells and normal cells following fluid-phase endocytosis.</p><p><b>RESULT</b>The nucleotide and amino acid sequences of the cloned MAP30 were identical with those of reported MAP30. The solubility of recombinant protein was analyzed by SDS-PAGE, and the MAP30 was mainly produced in soluble form. The recombinant MAP30 showed a greater cytotoxicity to cancer cells than that to normal cells.</p><p><b>CONCLUSION</b>The gene of MAP30 has been successfully cloned.The recombinant MAP30 protein expressed by E.coli is bioactive.</p>
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Seeds
/
Transformation, Bacterial
/
Recombinant Proteins
/
Gene Expression
/
Chemistry
/
Cloning, Molecular
/
Momordica charantia
/
Escherichia coli
/
Ribosome Inactivating Proteins, Type 2
/
Genetic Vectors
Language:
Chinese
Journal:
Journal of Zhejiang University. Medical sciences
Year:
2010
Type:
Article
Similar
MEDLINE
...
LILACS
LIS