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[目的 ]测定云南与海南两省不同疟区恶性疟原虫分离株谷氨酸富有蛋白 (GLURP)基因的部分序列及了解其分型。[方法 ]采用套式PCR方法特异性扩增GLURP基因R2区片段 ,并将该基因片段克隆于T载体 ,用双脱氧末端终止法测定不同长度的阳性克隆核苷酸序列 ,并应用DNAStar软件对云南与海南两省分离株GLURP基因及蛋白序列进行比较和分析。[结果 ]首次发现云南与海南两省恶性疟原虫至少存有 7个大小不同的GLURP等位基因型虫株 ,其基因片段变化范围为 6 0 0～ 15 0 0bp。不同分离株的GLURP基因R2区具高度保守性 ,其由编码 19～ 2 0个氨基酸的碱基的基本重复单位构成 ,该基因具有长度的多态性 ,表现在碱基基本重复单位的数目不同。序列分析结果表明 ,我国不同分离株之间或同一地区不同株之间GLURP基因及氨基酸序列具有高度的同源性 ,无明显的地理差异。 [结论 ]不同分离株GLURP基因结构的高度保守性及碱基重复片段数目的多态性 ,对研究疫苗候选抗原和建立疟原虫基因分型方法具有一定理论价值。
[Objective] To determine the partial sequence of GLURP gene of Plasmodium falciparum isolate in Yunnan and Hainan Provinces and to understand its classification. [Method] The R2 region fragment of GLURP gene was amplified by nested PCR method. The gene fragment was cloned into T vector. The positive cloned nucleotide sequences of different lengths were determined by dideoxy terminator method. The GLURP gene and protein sequences in Yunnan and Hainan provinces were compared and analyzed. [Results] For the first time, there were at least seven GLURP alleles with different sizes in P. falciparum in Yunnan and Hainan Provinces, and the range of their gene fragments varied from 600 to 1500 bp. The R2 region of the GLURP gene from different isolates is highly conserved and consists of a basic repeat unit encoding 19 to 20 amino acids with a length polymorphism expressed in different number of base repeat units . Sequence analysis showed that the GLURP gene and amino acid sequences among different isolates in China or in the same region had a high degree of homology with no obvious geographical differences. [Conclusion] The highly conserved structure of GLURP gene and the polymorphism of the number of repeated nucleotide fragments in different isolates have certain theoretical value for the study of vaccine candidate antigens and establishment of the genotyping method of Plasmodium.