DNA barcoding in Microorganisms. Alexandre Soares Rosado Institute of Microbiology UFRJ - Brazil. Bergey’s Manual = 4500 Species Some studies DNA reassociation= 10.000 genomes / g soil; 1% - 5% of microorganismos are culturable The majority = uncuturable .
Alexandre Soares Rosado
Institute of Microbiology
UFRJ - Brazil
Some studies DNA reassociation= 10.000 genomes / g soil;
1% - 5% of microorganismos are culturable
The majority = uncuturable
BRAZILIAN BIODIVERSITYFonte: Lewinsohn & Prado, 2000
composition and dynamics of soil microbial communities
Cultivation-independent analysis of large numbers
Sequencing of differentiating bands
Use of probes to identify bacterial isolates
corresponding to differentiating bands
Functionally conserved in all forms of life
The rRNA-gene - an ideal molecular marker?
„The molecular clock“
Regions of different degrees of conservation
Different number of ribosomal operons
A given variable region allows a different resolution for
Figure 1 the: gene rpoB
Figure 2: theDistance (p) among strains of Paenibacillus 16S and rpoB
PCR amplification of 16S
or 18S rDNA fragments
Few prominent populations (low evenness): patterns with few bands
Molecular fingerprints of
Denaturing gradient gel electrophoresis
Many equally abundant populations
patterns with many bands
Soil aggregation and bacterial community structure as theaffected by tillage and cover cropping in theBrazilian CerradosPeixoto et al., 2006
The Cerrados region in central Brazil occupies 22% of the country. It is characterized by high average temperature (22 - 270C), rainfall (800 - 1600 mm) and solar radiation (475- 500 Cal/cm2/day).
No Till X Till the
L TW1 TW2 T1 T2 NTW1 F2 NT1 NT2 F1 NTW2 L
Biodiversity F2 L
DGGE- Fingerprints 16S rDNA
Diversity of bacterial comunities
Identification/ 16S rDNA-Sequencing
1b Bacillus megaterium
2b Arthrobacter sp.
2s Streptomyces galbus
3s Streptomyces sp.
5s Pseudomonas sp.
1p Bacillus megaterium
2p unknown Bacterium
3p unknown Bacterium
1r Pseudomonas sp.
IV F2 L
Comparison between Denaturing Gradient Gel Electrophoresis (DGGE) and Phylogenetic Analysis for characterization of A/H3N2 Influenza Samples detected during 1999-2004 epidemics in Brazil.
Mangroves F2 L
Important issues F2 L
When the most efficient means of rapid barcode-based species identification is sought, a choice can be made either for one of these methodologies or for basic high-throughput sequencing, depending on the strategic outlook of the investigator and on current costs.
Arrays and functionally similar platforms may have a particular advantage when a biologically complex material such as soil or a human respiratory secretion sample is analysed to give a census of relevant species present.
Laboratory of Molecular Microbial Ecology –LMME