[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.
Gao
Yu-hui
YH
Etiology Department, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005, China.
Wang
Heng
H
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