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'''Ribotyping''' is a molecular technique for bacterial identification and characterization that uses information from rRNA-based phylogenetic analyses.<ref name=":0">{{Cite book|title=Biology of Microorganisms|last=Madigan|first=Michael T.|publisher=Pearson|year=2012|isbn=978-0-321-73551-5|pages=491}}</ref> It is a rapid and specific method widely used in clinical diagnostics and analysis of microbial communities in food, water, and beverages.<ref name=":0" /> |
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'''Ribotyping''' involves the [[Genetic fingerprinting|fingerprinting]] of genomic [[DNA]] [[Restriction digest|restriction fragments]] that contain all or part of the genes coding for the [[16S ribosomal RNA|16S]] and [[23S ribosomal RNA|23S]] [[rRNA]]. Conceptually, ribotyping is similar to probing restriction fragments of chromosomal DNA with [[Hybridization probe|cloned probes]] (randomly cloned probes or probes derived from a specific coding sequence such as that of a [[virulence factor]]). |
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All bacteria have [[ribosomal genes]], but the exact sequence is unique to each species, serving as a genetic fingerprint. Therefore, sequencing the particular 16S gene and comparing it to a database would yield identification of the particular species.<ref>{{Cite web|url=http://learn.genetics.utah.edu/content/gsl/diversity/|title=Investigating Microbial Diversity: Then and Now|website=learn.genetics.utah.edu|access-date=2016-03-15}}</ref> |
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==Technique== |
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Ribotyping involves the digestion of bacterial genomic DNA with specific [[restriction enzyme]]s. Each restriction enzyme cuts DNA at a specific nucleotide sequence, resulting in fragments of different lengths.<ref>{{Cite web|url=https://askabiologist.asu.edu/restriction-enzymes|title=Restriction Enzymes {{!}} ASU - Ask A Biologist|website=askabiologist.asu.edu|access-date=2016-03-13}}</ref> |
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Those fragments are then run on a [[Gel electrophoresis]], where they are separated according to size: the application of electrical field to the gel in which they are suspended causes the movement of DNA fragments (all negatively charged due to the presence of phosphate groups) through a matrix towards the positively charged end of the field. Small fragments move more easily and rapidly through the matrix, reaching a bigger distance from the starting position than larger fragments. |
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Following the separation in the gel matrix, the DNA fragments are moved onto nylon membranes and hybridized with a labelled 16S or 23S rRNA probe. This way only the fragments coding for such rRNA are visualised and can be analyzed.<ref>{{Cite web|url=https://courses.cit.cornell.edu/biomi290/microscopycases/methods/ribotype.htm|title=ribotype|website=courses.cit.cornell.edu|access-date=2016-03-13}}</ref> The pattern is then digitized and used to identify the origin of the DNA by a comparison with reference organisms in a computer database.<ref name=":0" /> |
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Conceptually, ribotyping is similar to probing restriction fragments of chromosomal DNA with [[Hybridization probe|cloned probes]] (randomly cloned probes or probes derived from a specific coding sequence such as that of a [[virulence factor]]).<ref>Grimont, F., and P. A. Grimont. 1986. Ribosomal ribonucleic acid gene restriction patterns as potential taxonomic tools. Ann. Inst. Pasteur Microbiol. 137B:165–175</ref> |
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==See also== |
==See also== |
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* [[Genotyping]] |
* [[Genotyping]] |
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==References== |
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[[it:Ribotipia]] |
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[[sv:Ribotypning]] |
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Ribotyping is a molecular technique for bacterial identification and characterization that uses information from rRNA-based phylogenetic analyses.[1] It is a rapid and specific method widely used in clinical diagnostics and analysis of microbial communities in food, water, and beverages.[1]
All bacteria have ribosomal genes, but the exact sequence is unique to each species, serving as a genetic fingerprint. Therefore, sequencing the particular 16S gene and comparing it to a database would yield identification of the particular species.[2]
Ribotyping involves the digestion of bacterial genomic DNA with specific restriction enzymes. Each restriction enzyme cuts DNA at a specific nucleotide sequence, resulting in fragments of different lengths.[3]
Those fragments are then run on a Gel electrophoresis, where they are separated according to size: the application of electrical field to the gel in which they are suspended causes the movement of DNA fragments (all negatively charged due to the presence of phosphate groups) through a matrix towards the positively charged end of the field. Small fragments move more easily and rapidly through the matrix, reaching a bigger distance from the starting position than larger fragments.
Following the separation in the gel matrix, the DNA fragments are moved onto nylon membranes and hybridized with a labelled 16S or 23S rRNA probe. This way only the fragments coding for such rRNA are visualised and can be analyzed.[4] The pattern is then digitized and used to identify the origin of the DNA by a comparison with reference organisms in a computer database.[1]
Conceptually, ribotyping is similar to probing restriction fragments of chromosomal DNA with cloned probes (randomly cloned probes or probes derived from a specific coding sequence such as that of a virulence factor).[5]
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