[Cloning and optimized prokaryotic expression of a pbmag-1 cDNA fragment from Plasmodium berghei ANKA].
OBJECTIVE
To clone and express a novel gene cDNA fragment, pbmag-1, from Plasmodium berghei ANKA strain.


METHODS
The cDNA sequence of pbmag-1 was obtained from the GenBank of P. berghei ANKA genomic databases, with which a pair of primers was designed and RT-PCR was used to get a cDNA fragment of the gene from the parasite. The expanded cDNA 3' fragment of the gene was obtained by 3'-RACE using the oligo dT primer and a set of specific primers. The intact cDNA 3' fragment was cloned into a prokaryotic expressional vector and transformed into the BL21-(DE3)-RIL strain of Escherichia coli. The recombinant protein of PbMAg-1 was expressed with an optimized strategy and used to immunize mice.


RESULTS
The pbmag-1 cDNA fragment obtained was 1341 bp in length, A/T rich (73%) and with a correct 3' end sequence. By Western blot, the anti-serum of mice immunized with the recombinant protein of PbMAg-1/GST, which was expressed as inclusion bodies, specifically recognized a band with Mr 64,000 molecule from the protein extracts of P. berghei-infected mouse erythrocytes.


CONCLUSIONS
The pbmag-l cDNA sequence with intact 3' has been obtained, which will be used for further study on its role in the immune response of P. berghei infection.
Etiology Department, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005, China.
Yu-hui
YH
Gao
Heng
H
Wang
D000818:Animals; D000913:Antibodies, Protozoan / Q000097:blood; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics; D005260:Female; D015870:Gene Expression; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010962:Plasmodium berghei / Q000235:genetics* / Q000276:immunology; D015800:Protozoan Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism
D000913:Antibodies, Protozoan; D018076:DNA, Complementary; D015800:Protozoan Proteins; D011994:Recombinant Proteins