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| Content Provider | World Health Organization (WHO)-Global Index Medicus |
|---|---|
| Author | Graças, D. A. Silva, A. Santos, A. S. Kodama, C. S. Cuadros-Orellana, S. Bandeira, C. H. M. M. |
| Description | Country affiliation: Brazil Author Affiliation: Kodama CS ( Laboratório de Polimorfismo de DNA, Instituto de Ciências Biológicas, Universidade Federal do Pará, Belém, PA, Brasil.); Cuadros-Orellana S ( Laboratório de Polimorfismo de DNA, Instituto de Ciências Biológicas, Universidade Federal do Pará, Belém, PA, Brasil.); Bandeira CH ( Laboratório de Investigação Sistemática em Biotecnologia, Instituto de Ciências Exatas e Naturais, Universidade Federal do Pará, Belém, PA, Brasil.); Graças DA ( Laboratório de Polimorfismo de DNA, Instituto de Ciências Biológicas, Universidade Federal do Pará, Belém, PA, Brasil.); Santos AS ( Laboratório de Investigação Sistemática em Biotecnologia, Instituto de Ciências Exatas e Naturais, Universidade Federal do Pará, Belém, PA, Brasil.); Silva A ( Laboratório de Polimorfismo de DNA, Instituto de Ciências Biológicas, Universidade Federal do Pará, Belém, PA, Brasil asilva@ufpa.br.) |
| Abstract | Denaturing high-performance liquid chromatography (DHPLC) has been described as a suitable method to study DNA polymorphisms. Here, cassava (Manihot esculenta Crantz) fermentation liquor was examined using DHPLC analysis to characterize the bacterial diversity during the fermentation process. GC-clamped amplicons corresponding to a variable region of the bacterial community 16S rDNA were synthesized using polymerase chain reaction (PCR) and then resolved on a base-composition basis using preparative DHPLC. Eluate fractions were collected at random and used as a source of whole community DNA that could be used to determine the bacterial diversity. As a first approach, GC-clamps were removed from the eluted DNA fragments using PCR to avoid the possible bias these clamps could cause during the construction of clone libraries. As a second approach, a clone library of each eluate sample was constructed, preserving the GC-clamps of the DNA fragments. The first approach generated 132 bacterial rDNA sequences with an average size of 200 bp, 45% of which had similarity to unculturable or non-classified bacteria. The second approach produced 194 sequences identified as Proteobacteria (48%), uncultured or non-classified environmental bacteria (40%) and Firmicutes (12%). We detected a remarkably greater bacterial diversity using the first approach than the second approach. The DHPLC-PCR method allowed for the fast and non-laborious detection of a vast bacterial diversity that was associated with cassava fermentation, and we conclude that it is a promising alternative for the characterization of the overall microbial diversity in complex samples. |
| e-ISSN | 16765680 |
| Journal | Genetics and Molecular Research |
| Issue Number | 1 |
| Volume Number | 13 |
| Language | English |
| Publisher | Fundação de Pesquisas Científicas de Ribeirão Preto |
| Publisher Date | 2014-02-28 |
| Publisher Place | Brazil |
| Access Restriction | Open |
| Subject Keyword | Bacteria Classification Isolation & Purification Manihot Microbiology Metagenome Genetics Chromatography, High Pressure Liquid Cloning, Molecular Colony Count, Microbial Dna, Bacterial Dna, Ribosomal Fermentation Microbiota Molecular Sequence Data Polymerase Chain Reaction Sequence Analysis, Dna Research Support, Non-u.s. Gov't Discipline Genetics Discipline Molecular Biology Discipline Bioinformatics |
| Content Type | Text |
| Resource Type | Article |
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