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- Title
Direct detection of mRNA expression in microbial cells by fluorescence in situ hybridization using RNase H-assisted rolling circle amplification.
- Authors
Takahashi, Hirokazu; Horio, Kyohei; Kato, Setsu; Kobori, Toshiro; Watanabe, Kenshi; Aki, Tsunehiro; Nakashimada, Yutaka; Okamura, Yoshiko
- Abstract
Meta-analyses using next generation sequencing is a powerful strategy for studying microbiota; however, it cannot clarify the role of individual microbes within microbiota. To know which cell expresses what gene is important for elucidation of the individual cell's function in microbiota. In this report, we developed novel fluorescence in situ hybridization (FISH) procedure using RNase-H-assisted rolling circle amplification to visualize mRNA of interest in microbial cells without reverse transcription. Our results show that this method is applicable to both Gram-negative and Gram-positive microbes without any noise from DNA, and it is possible to visualize the target mRNA expression directly at the single-cell level. Therefore, our procedure, when combined with data of meta-analyses, can help to understand the role of individual microbes in the microbiota.
- Subjects
MESSENGER RNA; GENE expression; MICROBIAL cells; FLUORESCENCE in situ hybridization; RIBONUCLEASE H; GENE amplification
- Publication
Scientific Reports, 2020, Vol 10, Issue 1, p1
- ISSN
2045-2322
- Publication type
Article
- DOI
10.1038/s41598-020-65864-7