Laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridization
Direct cultivation-independent sequence retrieval of unidentified bacteria from histological tissue sections has been limited by the difficulty of selectively isolating specific bacteria from a complex environment. Here, a new DNA isolation approach is presented for prokaryotic cells. By this method...
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| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Taylor & Francis Group
2005-12-01
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| Series: | BioTechniques |
| Online Access: | https://www.future-science.com/doi/10.2144/000112024 |
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| _version_ | 1850152914692079616 |
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| author | Kirstine Klitgaard Lars Mølbak Tim K. Jensen Christian Fredrik Lindboe Mette Boye |
| author_facet | Kirstine Klitgaard Lars Mølbak Tim K. Jensen Christian Fredrik Lindboe Mette Boye |
| author_sort | Kirstine Klitgaard |
| collection | DOAJ |
| description | Direct cultivation-independent sequence retrieval of unidentified bacteria from histological tissue sections has been limited by the difficulty of selectively isolating specific bacteria from a complex environment. Here, a new DNA isolation approach is presented for prokaryotic cells. By this method, a potentially pathogenic strain of the genus Brachyspira from formalin-fixed human colonic biopsies were visualized by fluorescence in situ hybridization (FISH) with a 16S rRNA-targeting oligonucleotide probe, followed by laser capture microdissection (LCM) of the targeted cells. Direct 16S rRNA gene PCR was performed from the dissected microcolonies, and the subsequent DNA sequence analysis identified the dissected bacterial cells as belonging to the Brachyspira aalborgi cluster 1. The advantage of this technique is the ability to combine the histological recognition of the specific bacteria within the tissue with molecular analysis of 16S rRNA gene or other genes of interest. This method is widely applicable for the identification of noncultivable bacteria and their gene pool from formalin-fixed paraffin-embedded tissue samples. |
| format | Article |
| id | doaj-art-77eacb79c7314da2acfbbde02b5bb656 |
| institution | OA Journals |
| issn | 0736-6205 1940-9818 |
| language | English |
| publishDate | 2005-12-01 |
| publisher | Taylor & Francis Group |
| record_format | Article |
| series | BioTechniques |
| spelling | doaj-art-77eacb79c7314da2acfbbde02b5bb6562025-08-20T02:25:51ZengTaylor & Francis GroupBioTechniques0736-62051940-98182005-12-0139686486810.2144/000112024Laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridizationKirstine Klitgaard0Lars Mølbak1Tim K. Jensen2Christian Fredrik Lindboe3Mette Boye41Danish Institute for Food and Veterinary Research, Copenhagen, Denmark1Danish Institute for Food and Veterinary Research, Copenhagen, Denmark1Danish Institute for Food and Veterinary Research, Copenhagen, Denmark2Sørlandet Sykehus HF, Kristiansand, Norway1Danish Institute for Food and Veterinary Research, Copenhagen, DenmarkDirect cultivation-independent sequence retrieval of unidentified bacteria from histological tissue sections has been limited by the difficulty of selectively isolating specific bacteria from a complex environment. Here, a new DNA isolation approach is presented for prokaryotic cells. By this method, a potentially pathogenic strain of the genus Brachyspira from formalin-fixed human colonic biopsies were visualized by fluorescence in situ hybridization (FISH) with a 16S rRNA-targeting oligonucleotide probe, followed by laser capture microdissection (LCM) of the targeted cells. Direct 16S rRNA gene PCR was performed from the dissected microcolonies, and the subsequent DNA sequence analysis identified the dissected bacterial cells as belonging to the Brachyspira aalborgi cluster 1. The advantage of this technique is the ability to combine the histological recognition of the specific bacteria within the tissue with molecular analysis of 16S rRNA gene or other genes of interest. This method is widely applicable for the identification of noncultivable bacteria and their gene pool from formalin-fixed paraffin-embedded tissue samples.https://www.future-science.com/doi/10.2144/000112024 |
| spellingShingle | Kirstine Klitgaard Lars Mølbak Tim K. Jensen Christian Fredrik Lindboe Mette Boye Laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridization BioTechniques |
| title | Laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridization |
| title_full | Laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridization |
| title_fullStr | Laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridization |
| title_full_unstemmed | Laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridization |
| title_short | Laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridization |
| title_sort | laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridization |
| url | https://www.future-science.com/doi/10.2144/000112024 |
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