| Project/Area Number |
07807030
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
寄生虫学(含医用動物学)
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| Research Institution | Kagawa Medical University |
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Principal Investigator |
SUGURI Setsuo Kagawa Medical University, Medical Zoology, Associate Professor, 医学部, 助教授 (00032897)
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| Co-Investigator(Kenkyū-buntansha) |
OKAMOTO Munehiro Osaka University, Experimental Animal Sciences, Research Assosiate, 医学部, 助手 (70177096)
HARADA Masakazu Kagawa Medical University, Medical Zoology, Research Associate, 医学部, 助手 (90127580)
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| Project Period (FY) |
1995 – 1996
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| Project Status |
Completed (Fiscal Year 1996)
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| Budget Amount *help |
¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1996: ¥300,000 (Direct Cost: ¥300,000)
Fiscal Year 1995: ¥1,600,000 (Direct Cost: ¥1,600,000)
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| Keywords | Parasite / identification / CO1 Gene / Nucleotide Sequence |
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| Research Abstract |
Segments of the CO1 (cytochrome c oxidase subunit 1) genes (391bp out of 1500bp) of Dirofilaria immitis, Paragonimus mexicanus and a sandfly were sequenced for the future examinations of species identification and phylogenic-tree making, and also compared with the sequences of some nematodes trematodes and insects.
Total DNAs were prepared from adult worms using phenol/chloroform extraction method. PCR amplifications were performed using a set of primers (5'TGG TTT TTT GTG CAT CCT GAG GTT TA3' and 5'AGA AAG AAC GTA ATG AAA ATG AGC AAC3') designed for the CO1 gene of planaria. The PCR products were cloned with pT7Blue T-Vector plasmid and NovaBlue competent cells (Novagen). The amplified plasmids were extracted with Wizard miniprep purification kit (Promega). Sequencing was done using a dye-primer cycle sequencing kit and a Model 377 DNA sequencer (Applied Biosystems). DNA sequence data were aligned and analyzed using the GENETYX computer program.
The nucleotide sequences of the PCR products were read from both ends and unambiguous sequences of 391bp were determined. The most nearly positive identical sequence of D.immitis was that of Ascaris suum in GeneBank nucleic acid sequence database and the identity was 71% (Poisson Probability ; 1.6e-80). The secondary identical sequence was that of Caenorhabditis elegans and the identity was 66%. In the case of P.mexicanus, the highest identity was shown with Fasciola hepatica (80%, PP ; 6.7e-92) and secondary highest identity with Schistosoma mekongi (76%, 4.0e-83). An unambiguous sequence of 388bp of a sandflies of Ecuador was also determined. The most nearly positive identical sequence of this was that of Drosophila melanogaster in GeneBank nucleic acid sequence database and the identity was 83% (Poisson Probability, 2.6e-102). The secondary identical sequence was that of Anopheles quadrimaculatus and the identity was also 83% ( Poisson Probability, 2.0e-105).
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