A modified in situ RT-PCR method for localizing fungal-specific gene expression in Candida-infected mice renal cells

Voon Chen Yong; Khang Wei Ong; Shiran Mohd Sidik; Rozita Rosli; Pei Pei Chong

doi:10.1016/j.mimet.2009.08.019

A modified in situ RT-PCR method for localizing fungal-specific gene expression in Candida-infected mice renal cells

J Microbiol Methods. 2009 Nov;79(2):242-5. doi: 10.1016/j.mimet.2009.08.019. Epub 2009 Sep 6.

Authors

Voon Chen Yong¹, Khang Wei Ong, Shiran Mohd Sidik, Rozita Rosli, Pei Pei Chong

Affiliation

¹ Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, University Putra Malaysia, Serdang, Selangor, Malaysia.

PMID: 19737582
DOI: 10.1016/j.mimet.2009.08.019

Abstract

In situ Reverse Transcriptase PCR (in situ RT-PCR) can amplify mRNA and localize gene expression in cells. However, this method is not feasible in fungi as the thick fungal cell wall constitutes a barrier to this procedure. We developed a two step in situ RT-PCR procedure which enabled the detection and localization of Candida tropicalis mRNA expression in formalin-fixed, paraffin-embedded (FFPE) mouse kidney sections. This in situ hybridization study revealed the first direct evidence for deposition of Candida tropicalis secreted aspartic proteinase 2 (CtSAP2) in the tip of pseudohyphae and its involvement in acute systemic candidiasis. We conclude that in situ RT-PCR can be successfully applied to FFPE tissues and will offer new perspectives in studying gene expression in Candida species.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Aspartic Acid Proteases / genetics
Candida tropicalis / genetics*
Candida tropicalis / pathogenicity
Candidiasis / microbiology*
Female
Gene Expression*
Genes, Fungal
In Situ Hybridization / methods*
Kidney / microbiology
Kidney Diseases / microbiology*
Mice
Mice, Inbred BALB C
Pathology / methods*
Reverse Transcriptase Polymerase Chain Reaction / methods*

Substances

Aspartic Acid Proteases