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summary
3
What you can expect to know:
0 Introduction
0 Different techniques of ribotyping
0 Adventages and limitations
0 Literature review
4
definition
Why rdna? it means…..
1.Universally present 2.large and stable 3.high
degree sequence conservation classification
Ribosomal RNA r DNA description-identification-nomenclature
ribotyping
Ribo … …typing
5
6
Organization of ribosomal operon
7
8
Conventional ribotyping
9
10
Selection of restriction endonuclease by sequence analysis(in silico)
11
Ribotyping prototype : Haemophilus influenzae strain Rd
In silico analysis
The presence of 6 ribosomal operon
Ribotyping protocol for Haemophilus
influenzae strain Rd
Stage 1-search the conserved restriction endonuclease site within the six
ribosomal operon.
Stage 2-choose the ideal restriction enzyme
Stage3-confirm the conservation of restriction site by the all publicity
available for 16-23s-rRNA sequencing
Stage4-cleavage electrophoresis southern blot hybridization with
a labelled probe containing conserved ribosomal sequences.
12
Cont.
0 The importance of this method :strain differentiation within the species
0 It implemented for several other bacterial species,eg : S.aureus
E.coli
p.aeroginosa
B.cepacia
N.meningitidis
B.pertussis
13
Automated ribotyping
0 Was first introduced by dupont qualicon-1995….. Riboprint pattern.
0 This system is reproducible ,convenient and fast
0 A single colony of bacteria is picked up and suspended in lysing buffer
and transferred to the riboprinter in the riboprinter like
conventional ribotyping….
14
PCR Ribotyping
15
Advantages and disadvantages
Advantages of ribotyping as an identification method:
0 Ribotyping is a fully automated procedure
0 Procedure involves less labor and is standardized
Disadvantages of ribotyping:
0 Expensive because of the equipment used, therefore usually
only performed in reference laboratories
Literature review
references
• Grimont, F., and P. A. D. Grimont. 1986. Ribosomal ribonucleic acid
gene restriction patterns aspotential taxonomic
tools. Ann. Inst. Pasteur Micro- biol.
• Altwegg, M., F. W. Hickman-Brenner, and J. J. Farmer III. 1989.
Ribosomal RNA gene restriction patterns provide increased sensitivity
for typing Salmonella typhi strains. J. Infect. Dis. 160:145–149.
• Bingen, E. H., P. Mariani-Kurkdjian, N. Y. Lambert-Zechovsky, P.
Dejardins, E. Denamur, Y. Aujard, E. Vilmer, and J. Elion. 1992.
Ribotyping provides efficient differentiation of nosocomial Serratia
marcescens isolates in a pediatric hospital. J. Clin. Microbiol.
30:2088–2091.
Cont.
Farmer, J. J., III. 1972. Epidemiological differentiation of Serratia marcescens:
typing by bacteriocin sensitivity. Appl. Microbiol. 23:226–231.
• Animal Biotechnology: Models in Discovery and Translationedited by
Ashish Verma, Anchal Singh
24
25

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Ribotyping

  • 1.
  • 2.
  • 4. What you can expect to know: 0 Introduction 0 Different techniques of ribotyping 0 Adventages and limitations 0 Literature review 4
  • 5. definition Why rdna? it means….. 1.Universally present 2.large and stable 3.high degree sequence conservation classification Ribosomal RNA r DNA description-identification-nomenclature ribotyping Ribo … …typing 5
  • 6. 6
  • 8. 8
  • 10. 10
  • 11. Selection of restriction endonuclease by sequence analysis(in silico) 11 Ribotyping prototype : Haemophilus influenzae strain Rd In silico analysis The presence of 6 ribosomal operon
  • 12. Ribotyping protocol for Haemophilus influenzae strain Rd Stage 1-search the conserved restriction endonuclease site within the six ribosomal operon. Stage 2-choose the ideal restriction enzyme Stage3-confirm the conservation of restriction site by the all publicity available for 16-23s-rRNA sequencing Stage4-cleavage electrophoresis southern blot hybridization with a labelled probe containing conserved ribosomal sequences. 12
  • 13. Cont. 0 The importance of this method :strain differentiation within the species 0 It implemented for several other bacterial species,eg : S.aureus E.coli p.aeroginosa B.cepacia N.meningitidis B.pertussis 13
  • 14. Automated ribotyping 0 Was first introduced by dupont qualicon-1995….. Riboprint pattern. 0 This system is reproducible ,convenient and fast 0 A single colony of bacteria is picked up and suspended in lysing buffer and transferred to the riboprinter in the riboprinter like conventional ribotyping…. 14
  • 16. Advantages and disadvantages Advantages of ribotyping as an identification method: 0 Ribotyping is a fully automated procedure 0 Procedure involves less labor and is standardized Disadvantages of ribotyping: 0 Expensive because of the equipment used, therefore usually only performed in reference laboratories
  • 18.
  • 19.
  • 20.
  • 21.
  • 22. references • Grimont, F., and P. A. D. Grimont. 1986. Ribosomal ribonucleic acid gene restriction patterns aspotential taxonomic tools. Ann. Inst. Pasteur Micro- biol. • Altwegg, M., F. W. Hickman-Brenner, and J. J. Farmer III. 1989. Ribosomal RNA gene restriction patterns provide increased sensitivity for typing Salmonella typhi strains. J. Infect. Dis. 160:145–149. • Bingen, E. H., P. Mariani-Kurkdjian, N. Y. Lambert-Zechovsky, P. Dejardins, E. Denamur, Y. Aujard, E. Vilmer, and J. Elion. 1992. Ribotyping provides efficient differentiation of nosocomial Serratia marcescens isolates in a pediatric hospital. J. Clin. Microbiol. 30:2088–2091.
  • 23. Cont. Farmer, J. J., III. 1972. Epidemiological differentiation of Serratia marcescens: typing by bacteriocin sensitivity. Appl. Microbiol. 23:226–231. • Animal Biotechnology: Models in Discovery and Translationedited by Ashish Verma, Anchal Singh
  • 24. 24
  • 25. 25