Subcloning and Sequencing of Conserved Blocks (Ⅱ 、 Ⅲ、Ⅳ) of P190 Gene of Plasmodium falciparum FCC1/HN Isolate / 第二军医大学学报

ABSTRACT

190-kilodalton glycoprotein (P190) of Plasmodium falciparum. precursor of the major surface protein of merozoites, is considered a promising candidate for blood stage malarial vaccine. We designed six primers according to the sequence of MAD20 strain, with a GC clamp and BamHI site at the 5'- end of each one, and a GC clamp and Xbal site at the 3'- end of each one. The primers were synthesized by phosphoramidite approach (User's Manual of ABI Company) and purified using HPLC. Three fragments in the second, third and fourth conserved regions of P190 gene of Plasmodium falciparum FCC1/HN strain isolated from the blood of patients in Hainan Province of China were amplified by the polymerase chain reaction (PCR) technique. The amplified fragments were subcloned into pUC18 vectors and sequenced using the dideoxy chain termination method. All three regions of P190 gene of FCCl/HN strain also were highly conservative as compared with P190 gene of MAD20 (Papua New Guinea isolate), K1 (Thailand isolate), Wellcome (West Africa isolate) and CAMP (Malaysia) strains of Plasmodium falciparum. The C at position 81 in the second conserved block of P190 gene of FCC1/HN isolate was substituted by T, which did not change the amino acid determined by the coden corresponding to the substitution.