A single-tube nested PCR for Pneumocystis carinii f. sp. hominis

Xing Tang, Marilyn S. Bartlett, James W. Smith, Chao-Hung Lee

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

A single-tube nested PCR which amplifies the internal transcribed spacer (ITS) regions of the rRNA genes of human Pneumocystis carinii was developed. The outer primers for the first PCR, which anneal to the 18S and the 26S rRNA genes of P. carinii, were made to have a midpoint temperature (T(m) of 74°C. The inner primers for the second PCR have a T(m) of 56 to 58°C and are specific for human P. carinii; they anneal to an area close to the beginning of ITS1 and the junction of ITS2 and the 26S rRNA genes. The reaction mixture contained 2.5 pmol of the first-PCR primers and 25 pmol of the second-PCR primers. The first PCR was performed at an annealing temperature of 68°C, which did not allow the second-PCR primers to function. Since very small amounts (2.5 pmol) of the first-PCR primers were used, they were exhausted when the first PCR was completed. The single-tube nested PCR did not amplify P. carinii isolated from rats, mice, or ferrets. All 10 bronchoalveolar lavage (BAL) specimens from patients with P. carinii pneumonia were positive, whereas all 10 BAL specimens from patients with other diseases or patients infected with several commonly found fungi were negative by PCR.

Original languageEnglish
Pages (from-to)1597-1599
Number of pages3
JournalJournal of Clinical Microbiology
Volume35
Issue number6
StatePublished - Jun 1997

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Pneumocystis carinii
Polymerase Chain Reaction
rRNA Genes
Bronchoalveolar Lavage
Ferrets
Temperature
Pneumocystis Pneumonia
Fungi

ASJC Scopus subject areas

  • Microbiology (medical)
  • Microbiology

Cite this

A single-tube nested PCR for Pneumocystis carinii f. sp. hominis. / Tang, Xing; Bartlett, Marilyn S.; Smith, James W.; Lee, Chao-Hung.

In: Journal of Clinical Microbiology, Vol. 35, No. 6, 06.1997, p. 1597-1599.

Research output: Contribution to journalArticle

Tang, X, Bartlett, MS, Smith, JW & Lee, C-H 1997, 'A single-tube nested PCR for Pneumocystis carinii f. sp. hominis', Journal of Clinical Microbiology, vol. 35, no. 6, pp. 1597-1599.
Tang, Xing ; Bartlett, Marilyn S. ; Smith, James W. ; Lee, Chao-Hung. / A single-tube nested PCR for Pneumocystis carinii f. sp. hominis. In: Journal of Clinical Microbiology. 1997 ; Vol. 35, No. 6. pp. 1597-1599.
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