title: Cycle DNA sequencing with [alpha-35S]dATP demonstrates polymorphism of a surface antigen in malaria parasites from Sri Lankan patients.
abstract: Structural diversity in a 45 kDa surface antigen on Plasmodium falciparum merozoites (termed GYMSSA, MSP-2 or MSA-2) and other candidate molecules for developing a malaria vaccine need to be investigated in parasites obtained directly from patients in different malaria endemic countries. A double-stranded DNA sequencing method suitable for this purpose, and also for studying diversity in genes of other haploid cells, is described. A first round polymerase chain reaction (PCR) on DNA isolated from blood was carried out with a primer containing the GCN4 binding site to amplify and subsequently purify the coding region of the MSA-2 gene on GCN4 coated tubes. A second round PCR with more internal primers incorporating M13 forward and reverse primer sequences was then performed. Cycle sequencing was done with unlabelled M13 primers and [alpha-35S]dATP by the dideoxynucleotide procedure. Two different allelic forms of MSA-2 were identified in samples of Plasmodium falciparum from patients in Sri Lanka.
authors.affiliation: Division of Life Sciences, Institute of Fundamental Studies, Kandy, Sri Lanka.
authors.forename: R
authors.initials: R
authors.lastname: Ramasamy
authors.forename: C
authors.initials: C
authors.lastname: Ranasinghe
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000953:Antigens, Protozoan / Q000235:genetics*; D000954:Antigens, Surface / Q000235:genetics*; D001483:Base Sequence; D018076:DNA, Complementary / Q000032:analysis* / Q000302:isolation & purification; D003838:Deoxyadenine Nucleotides / Q000032:analysis; D006801:Humans; D016778:Malaria, Falciparum / Q000097:blood / Q000453:epidemiology; D008562:Membrane Glycoproteins / Q000235:genetics*; D008969:Molecular Sequence Data; D010963:Plasmodium falciparum / Q000235:genetics* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods; D011110:Polymorphism, Genetic; D015800:Protozoan Proteins; D013188:Sri Lanka
chemical_list: D000953:Antigens, Protozoan; D000954:Antigens, Surface; D018076:DNA, Complementary; D003838:Deoxyadenine Nucleotides; C064417:GYMSSA antigen, Plasmodium falciparum; D008562:Membrane Glycoproteins; D015800:Protozoan Proteins