Visualization of Gene Reciprocity among Lactic Acid Bacteria in Yogurt by RNase H-Assisted Rolling Circle Amplification-Fluorescence <i>In Situ</i> Hybridization
Recently, we developed an <i>in situ</i> mRNA detection method termed RNase H-assisted rolling circle amplification-fluorescence <i>in situ</i> hybridization (RHa-RCA-FISH), which can detect even short mRNA in a bacterial cell. However, because this FISH method is sensitive t...
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MDPI AG
2021-06-01
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| Series: | Microorganisms |
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| Online Access: | https://www.mdpi.com/2076-2607/9/6/1208 |
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| author | Kyohei Horio Hirokazu Takahashi Toshiro Kobori Kenshi Watanabe Tsunehiro Aki Yutaka Nakashimada Yoshiko Okamura |
| author_facet | Kyohei Horio Hirokazu Takahashi Toshiro Kobori Kenshi Watanabe Tsunehiro Aki Yutaka Nakashimada Yoshiko Okamura |
| author_sort | Kyohei Horio |
| collection | DOAJ |
| description | Recently, we developed an <i>in situ</i> mRNA detection method termed RNase H-assisted rolling circle amplification-fluorescence <i>in situ</i> hybridization (RHa-RCA-FISH), which can detect even short mRNA in a bacterial cell. However, because this FISH method is sensitive to the sample condition, it is necessary to find a suitable cell permeabilization and collection protocol. Here, we demonstrate its further applicability for detecting intrinsic mRNA expression using lactic acid bacteria (LAB) as a model consortium. Our results show that this method can visualize functional gene expression in LAB cells and can be used for monitoring the temporal transition of gene expression. In addition, we also confirmed that data obtained from bulk analyses such as RNA-seq or microarray do not always correspond to gene expression in individual cells. RHa-RCA-FISH will be a powerful tool to compensate for insufficient data from metatranscriptome analyses while clarifying the carriers of function in microbial consortia. By extending this technique to capture spatiotemporal microbial gene expression at the single-cell level, it will be able to characterize microbial interactions in phytoplankton–bacteria interactions. |
| first_indexed | 2024-03-10T10:45:13Z |
| format | Article |
| id | doaj.art-c502bd22e4b04cf6b5ecc0552a2981da |
| institution | Directory Open Access Journal |
| issn | 2076-2607 |
| language | English |
| last_indexed | 2024-03-10T10:45:13Z |
| publishDate | 2021-06-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Microorganisms |
| spelling | doaj.art-c502bd22e4b04cf6b5ecc0552a2981da2023-11-21T22:39:15ZengMDPI AGMicroorganisms2076-26072021-06-0196120810.3390/microorganisms9061208Visualization of Gene Reciprocity among Lactic Acid Bacteria in Yogurt by RNase H-Assisted Rolling Circle Amplification-Fluorescence <i>In Situ</i> HybridizationKyohei Horio0Hirokazu Takahashi1Toshiro Kobori2Kenshi Watanabe3Tsunehiro Aki4Yutaka Nakashimada5Yoshiko Okamura6Graduate School of Integrated Sciences for Life, Hiroshima University, Higashihiroshima 739-8530, JapanGraduate School of Integrated Sciences for Life, Hiroshima University, Higashihiroshima 739-8530, JapanDivision of Food Biotechnology, Food Research Institute, National Agriculture and Food Research Organization, Tsukuba, Ibaraki 305-8642, JapanGraduate School of Integrated Sciences for Life, Hiroshima University, Higashihiroshima 739-8530, JapanGraduate School of Integrated Sciences for Life, Hiroshima University, Higashihiroshima 739-8530, JapanGraduate School of Integrated Sciences for Life, Hiroshima University, Higashihiroshima 739-8530, JapanGraduate School of Integrated Sciences for Life, Hiroshima University, Higashihiroshima 739-8530, JapanRecently, we developed an <i>in situ</i> mRNA detection method termed RNase H-assisted rolling circle amplification-fluorescence <i>in situ</i> hybridization (RHa-RCA-FISH), which can detect even short mRNA in a bacterial cell. However, because this FISH method is sensitive to the sample condition, it is necessary to find a suitable cell permeabilization and collection protocol. Here, we demonstrate its further applicability for detecting intrinsic mRNA expression using lactic acid bacteria (LAB) as a model consortium. Our results show that this method can visualize functional gene expression in LAB cells and can be used for monitoring the temporal transition of gene expression. In addition, we also confirmed that data obtained from bulk analyses such as RNA-seq or microarray do not always correspond to gene expression in individual cells. RHa-RCA-FISH will be a powerful tool to compensate for insufficient data from metatranscriptome analyses while clarifying the carriers of function in microbial consortia. By extending this technique to capture spatiotemporal microbial gene expression at the single-cell level, it will be able to characterize microbial interactions in phytoplankton–bacteria interactions.https://www.mdpi.com/2076-2607/9/6/1208mRNAfluorescence <i>in situ</i> hybridizationlactic acid bacteriasingle cellsymbioses |
| spellingShingle | Kyohei Horio Hirokazu Takahashi Toshiro Kobori Kenshi Watanabe Tsunehiro Aki Yutaka Nakashimada Yoshiko Okamura Visualization of Gene Reciprocity among Lactic Acid Bacteria in Yogurt by RNase H-Assisted Rolling Circle Amplification-Fluorescence <i>In Situ</i> Hybridization Microorganisms mRNA fluorescence <i>in situ</i> hybridization lactic acid bacteria single cell symbioses |
| title | Visualization of Gene Reciprocity among Lactic Acid Bacteria in Yogurt by RNase H-Assisted Rolling Circle Amplification-Fluorescence <i>In Situ</i> Hybridization |
| title_full | Visualization of Gene Reciprocity among Lactic Acid Bacteria in Yogurt by RNase H-Assisted Rolling Circle Amplification-Fluorescence <i>In Situ</i> Hybridization |
| title_fullStr | Visualization of Gene Reciprocity among Lactic Acid Bacteria in Yogurt by RNase H-Assisted Rolling Circle Amplification-Fluorescence <i>In Situ</i> Hybridization |
| title_full_unstemmed | Visualization of Gene Reciprocity among Lactic Acid Bacteria in Yogurt by RNase H-Assisted Rolling Circle Amplification-Fluorescence <i>In Situ</i> Hybridization |
| title_short | Visualization of Gene Reciprocity among Lactic Acid Bacteria in Yogurt by RNase H-Assisted Rolling Circle Amplification-Fluorescence <i>In Situ</i> Hybridization |
| title_sort | visualization of gene reciprocity among lactic acid bacteria in yogurt by rnase h assisted rolling circle amplification fluorescence i in situ i hybridization |
| topic | mRNA fluorescence <i>in situ</i> hybridization lactic acid bacteria single cell symbioses |
| url | https://www.mdpi.com/2076-2607/9/6/1208 |
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