›› 1998, Vol. 16 ›› Issue (6): 406-410.

• 论著 • Previous Articles     Next Articles

CLONING AND SEQUENCING OF THE GENE CODING THE SEXUAL STAGE ANTIGEN Pfs48/45 OF PLASMODIUM FALCIPARUM

Luo Shuhong; Yu Xinbing; Liu Yanwen; Fang Jianming; Xu Jing; Li Xuerong   

  1. Department of Parasitology; Sun Yat sen University of MedicalSciences; Guangzhou 510089
  • Received:1900-01-01 Revised:1900-01-01 Online:1998-12-31 Published:1998-12-31

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Abstract: AIM: To express the antigen Pfs48/45 in vitro and provide an antigen for the
development of the transmission blocking vaccine. METHODS:According to the published nucleotide
sequence of Pfs48/45 of Plasmodium falciparum isolate NF54, a pair of oligonucleotides was
designed and used as primers(P1,P2). The gene encoding the gametocyte/gamete specific membrane
protein Pfs48/45 of P. falciparum isolate FCC1/HN has been amplified by using polymerase chain
reaction(PCR) technique. The PCR product was purified and directly sequenced by the dideoxynucleotide terminator method with 5’-end primer P1. At the same time, the purified PCR product was digested with BamHI and EcoRI and cloned into the plasmid pcDNA 3, then the recombinant clones were transformed into E. coli strain TG1. The recombinant plasmid pcDNA 32Pfs48/45 was screened and identified by PCR amplification and restriction analysis. RESULTS: ① The gene fragment Pfs48/45 was specifically amplified from the genomicDNA of Plasmodium falciparum isolate FCC1/HN; ②The sequence demon-strated that the 5’-end nucleotide and predicted aminoacid sequence of Pfs48/45 from FCC1/HN isolate was basically identical with that from NF54 isolate. We found that the sequence of the Pfs48/45 gene from the isolate FCC1/HN differs from the published sequence (isolate NF54) only at positions 307 and 372 (T →C). The substitution of T →C at the position of 372 generates a new restriction site Taq I. The PCR product digested by Taq I generates DNA fragment of 984 bp and 379 bp , suggesting that the PCR product is the gene encoding the transmission blocking antigen Pfs48/45; ③ The gene fragment of
Pfs48/45 was directly inserted into the BamHI and EcoRI site of plasmid pcDNA 3. CONCLUSION: The nucleotide sequence of Pfs48/4 5 of Plasmodium falciparum isolate FCC1/HN from south China was similar to that of isolate NF54. The recombinant plasmid pcDNA 3 Pfs48/45 was successfully constructed, providing a means to evaluate the role and biological function of this sexual-stage-specific protein of Pfs48/45.

Key words: Plasmodium falciparum, transmission blocking immunity, clone DNA sequence analysis, malaria vaccine