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The isolation of differentially expressed cDNA clones from the filarial nematode Brugia pahangi

Published online by Cambridge University Press:  01 August 1999

S. J. HUNTER ,
S. A. M. MARTIN ,
F. J. THOMPSON ,
L. TETLEY  and
E. DEVANEY
S. J. HUNTER
Affiliation:
Department of Veterinary Parasitology, University of Glasgow, Bearsden Road, Glasgow G61 1QH
S. A. M. MARTIN
Affiliation:
Department of Veterinary Parasitology, University of Glasgow, Bearsden Road, Glasgow G61 1QH
F. J. THOMPSON
Affiliation:
Department of Veterinary Parasitology, University of Glasgow, Bearsden Road, Glasgow G61 1QH
L. TETLEY
Affiliation:
Integrated Microscopy Facility, Division of Infection and Immunity, Joseph Black Building, University of Glasgow
E. DEVANEY
Affiliation:
Department of Veterinary Parasitology, University of Glasgow, Bearsden Road, Glasgow G61 1QH
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Abstract

A cDNA library constructed from 3 day post-infective L3 of the filarial nematode Brugia pahangi was screened by differential hybridization with cDNA probes prepared from different life-cycle stages. Five cDNA clones hybridizing selectively to the mosquito-derived L3 probe were isolated and characterized. Northern blot analysis of 4 of the clones confirmed that each was most highly expressed in the mosquito-derived L3. The expression of each mRNAduring parasite development in the mosquito vector was investigated using RT–PCR, and all were shown to be abundant in the immature L3. Four of the 5 cDNAs cloned coded for structural proteins: 2 cuticular collagens, and the muscle proteins tropomyosin and troponin. Further studies on troponin using an antiserum raised to the recombinant protein demonstrated that the protein, unlike the mRNA, was present in all life-cycle stages examined, while immunogold labelling demonstrated that it was localized to the muscle blocks.

Keywords

Brugia pahangiL3differentially expressed genestroponintropomyosincuticular collagens
Type
Research Article
Information
Parasitology , Volume 119 , Issue 2 , August 1999 , pp. 189 - 198
Copyright
1999 Cambridge University Press

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Footnotes

Nucleotide sequence data reported in this paper are available in the EMBL and GenBankTM data bases under the accession numbers: SJ1 AJ224966; SJ5 AJ224967; clone A AJ130821; clone C AJ224968; clone F AJ224969; clone EE4 X91066.