SPÓŁCZESNE METODY IDENTYFIKACJI
BAKTERII STOSOWANE W EKOLOGII
MIKROORGANIZMÓW WODNYCH
- FLUORESCENCYJNA HYBRYDYZACJA IN SITU (FISH)
Agnieszka Skowrońska, Izabella Zmysłowska
Uniwersytet Warmińsko-Mazurski, Katedra Mikrobiologii Środowiskowej
ul. Romana Prawocheńskiego 1, 10-957 Olsztyn-Kortowo
e-mail: agnieszka.skowronska@uwm.edu.pl
Wpłynęło w sierpniu 2005 r.
1. Wstęp. 2. Badania zespołów mikroorganizmów z zastosowaniem metody PCR i sekwencjonowaniem genów
rRNA. 3. Fluorescencyjna hybrydyzacja in situ (FISH). 4. Enzymatyczne wzmocnienie sygnału (CARD)-FISH. 5. Ograniczenia związane ze stosowaniem metody FISH. 6. Podsumowanie
Current methods for identification of bacteria in aquatic microbial ecology - fluorescence
in situ hybridization (FISH)
Abstract: Nowadays using molecular techniques prokaryotic cells can be quickly and directly identified in environmental samples. Fluorescence
in situ hybridization (FISH) with rRNA-targeted oligonucleotide probes has become one of the major techniques in environmental microbiology allows rapid and reliable quantification of population sizes, as well as distinction of bacterial morphotypes. The major question in microbial ecology of who is out there can be answered using this method. With the increased sensitivity of FISH and methodological improvements, as well as combination of FISH with various other techniques, more detailed account of the activity of a single microbial cell can be also gained. Another approach in microbial ecology is based on comparative sequence analysis of rRNA genes for the study of diversity and community composition of microbial assemblages. It has been termed the "full-cycle rRNA analysis" and offers a cultivation-independent alternative technique for the identification of new micro-organisms. In this procedure the 16S rRNA gene fragments can be selectively amplified from mixed DNA then cloned and sequenced. The rapidly growing amount of new retrieved seąuence allows comparative sequence analysis with the available databases and design of specific rRNA-targeted DNA probes.
1. Introduction. 2. Bacterial community analysis using PCR and sequencing of rRNA genes. 3. Fluorescence
in situ hybridization (FISH). 4. Enzymatic signal amplification (CARD)-FISH. 5. Current obstacles to general application of FISH method. 6. Summary
Słowa kluczowe: FISH, hybrydyzacja in situ, 16S rRNA, sondy
oligonukleotydowe
Key words: FISH, in situ hybridization, 16S rRNA, oligonucleotide probes
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