title: Haplorchis taichui, Witenberg, 1930: Development of a HAT-RAPD marker for the detection of minute intestinal fluke infection.
abstract: Specific primers to determine the presence of an intestinal fluke, Haplorchis taichui, were investigated using the high annealing temperature random amplified polymorphic DNA (HAT-RAPD) PCR, and 18 arbitrary primers (Operon Technologies), to generate different polymorphic DNA profiles. Thirteen kinds of parasites were used to compare fingerprints. A 256bp HAT-RAPD marker, generated from the OPP-11 primer, was found to be H. taichui-specific, and this marker was cloned, transformed, and sequenced. From the sequence data, a pair of primers were designed with Genetyx-MAC ver.11 and indicated as: Hap-t F 5'-GGC CAA CGC AAT CGT CAT CC-3' and Hap-t R 5'-GCG TCG GGT TTC AGA CAT GG-3'. These specific primers were tested for efficacy and specificity by amplifying them with all 13 parasites DNAs in PCR reaction. A 256bp amplicon was generated, which was shown to have a positive result, only for H. taichui DNA. It revealed no cross-reaction with any of the other tested parasite species. The minimum DNA template, needed for detection by PCR, was 0.1picogram (pg). The successful development of H. taichui-specific primers is expected to be beneficial for epidemiological studies and for prevention and control of these parasitic infections.
authors: Department of Biology, Chiang Mai University, Chiang Mai Province, Thailand. wchalobol@yahoo.com
authors: Chalobol
authors: C
authors: Wongsawad
authors: Pheravut
authors: P
authors: Wongsawad
authors: Jong Yil
authors: JY
authors: Chai
authors: Somboon
authors: S
authors: Anuntalabhochai
mesh_terms: D000818:Animals; D001483:Base Sequence; D002417:Cattle; D002645:Chickens; D006224:Cricetinae; D016172:DNA Fingerprinting; D017931:DNA Primers / Q000592:standards; D018509:DNA, Helminth / Q000737:chemistry / Q000302:isolation & purification*; D005399:Fishes; D006578:Heterophyidae / Q000235:genetics / Q000302:isolation & purification*; D007411:Intestinal Diseases, Parasitic / Q000175:diagnosis* / Q000469:parasitology; D008969:Molecular Sequence Data; D019105:Random Amplified Polymorphic DNA Technique; D011898:Ranidae; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D013045:Species Specificity; D014201:Trematode Infections / Q000175:diagnosis* / Q000469:parasitology
chemical_list: D017931:DNA Primers; D018509:DNA, Helminth