Amplification of a Cryptosporidium parvum gene fragment encoding thymidylate synthase.

L. Goozé, K. Kim, C. Petersen, J. Gut, R. G. Nelson

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Currently, there is no effective therapy for cryptosporidiosis and it is unclear why antifolate drugs which are effective treatments for infections caused by closely related parasites are not also effective against Cryptosporidium parvum. In protozoa, the target of these drugs, dihydrofolate reductase (DHFR), exists as a bifunctional enzyme also manifesting thymidylate synthase (TS) activity and is encoded by a fused DHFR-TS gene. In order to prepare a probe to isolate the C. parvum DHFR-TS gene we have used degenerate oligonucleotides whose sequences are based on strongly conserved regions of TS protein sequence to prime the polymerase chain reaction (PCR) with C. parvum DNA. The PCR amplified a 375-bp DNA fragment which was cloned and sequenced; the deduced amino acid sequence had significant identity with known TS sequences, including strict conservation of all phylogenetically invariant TS amino acid residues. The cloned PCR fragment was used as a probe to isolate a number of overlapping clones from a C. parvum genomic library which were definitively shown to be of cryptosporidial origin by genomic Southern and molecular karyotype analyses. The deduced protein sequence of C. parvum TS was most similar to the bifunctional TS enzymes of Plasmodium chabaudi and Plasmodium falciparum.

Original languageEnglish (US)
JournalJournal of Protozoology
Volume38
Issue number6
StatePublished - Nov 1991
Externally publishedYes

Fingerprint

Cryptosporidium parvum
Thymidylate Synthase
Genes
Polymerase Chain Reaction
Plasmodium chabaudi
Folic Acid Antagonists
Cryptosporidiosis
Tetrahydrofolate Dehydrogenase
Genomic Library
DNA
Enzymes
Plasmodium falciparum
Protein C
Karyotype
Oligonucleotides
Pharmaceutical Preparations
Amino Acid Sequence
Parasites
Clone Cells
Amino Acids

ASJC Scopus subject areas

  • Parasitology

Cite this

Goozé, L., Kim, K., Petersen, C., Gut, J., & Nelson, R. G. (1991). Amplification of a Cryptosporidium parvum gene fragment encoding thymidylate synthase. Journal of Protozoology, 38(6).

Amplification of a Cryptosporidium parvum gene fragment encoding thymidylate synthase. / Goozé, L.; Kim, K.; Petersen, C.; Gut, J.; Nelson, R. G.

In: Journal of Protozoology, Vol. 38, No. 6, 11.1991.

Research output: Contribution to journalArticle

Goozé, L, Kim, K, Petersen, C, Gut, J & Nelson, RG 1991, 'Amplification of a Cryptosporidium parvum gene fragment encoding thymidylate synthase.', Journal of Protozoology, vol. 38, no. 6.
Goozé, L. ; Kim, K. ; Petersen, C. ; Gut, J. ; Nelson, R. G. / Amplification of a Cryptosporidium parvum gene fragment encoding thymidylate synthase. In: Journal of Protozoology. 1991 ; Vol. 38, No. 6.
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