Efficiency of boiling and four other methods for genomic DNA extraction of deteriorating spore-forming bacteria from milk
| Autor: | Ronaldo Tamanini, Fernando de Godoi Silva, Bruna Fritegoto Soares, Francine Fernandes da Silva, José Carlos Ribeiro Júnior, Nayara Assis Augusto, Vanerli Beloti, Aline Marangon de Oliveira |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Lysis Microorganism Bacillus Microbiology law.invention Spores 03 medical and health sciences chemistry.chemical_compound law Food science lcsh:Agriculture (General) Polymerase chain reaction biology Raw milk biology.organism_classification DNA extraction lcsh:S1-972 genomic DNA 030104 developmental biology PCR chemistry General Agricultural and Biological Sciences Paenibacillus Bacteria DNA |
| Zdroj: | Semina: Ciências Agrárias, Vol 37, Iss 5, Pp 3069-3078 (2016) Redalyc |
| ISSN: | 1679-0359 |
| Popis: | The spore-forming microbiota is mainly responsible for the deterioration of pasteurized milk with long shelf life in the United States. The identification of these microorganisms, using molecular tools, is of particular importance for the maintenance of the quality of milk. However, these molecular techniques are not only costly but also labor-intensive and time-consuming. The aim of this study was to compare the efficiency of boiling in conjunction with four other methods for the genomic DNA extraction of sporulated bacteria with proteolytic and lipolytic potential isolated from raw milk in the states of Paraná and Maranhão, Brazil. Protocols based on cellular lysis by enzymatic digestion, phenolic extraction, microwave-heating, as well as the use of guanidine isothiocyanate were used. This study proposes a method involving simple boiling for the extraction of genomic DNA from these microorganisms. Variations in the quality and yield of the extracted DNA among these methods were observed. However, both the cell lysis protocol by enzymatic digestion (commercial kit) and the simple boiling method proposed in this study yielded sufficient DNA for successfully carrying out the Polymerase Chain Reaction (PCR) of the rpoB and 16S rRNA genes for all 11 strains of microorganisms tested. Other protocols failed to yield sufficient quantity and quality of DNA from all microorganisms tested, since only a few strains have showed positive results by PCR, thereby hindering the search for new microorganisms. Thus, the simple boiling method for DNA extraction from sporulated bacteria in spoiled milk showed the same efficacy as that of the commercial kit. Moreover, the method is inexpensive, easy to perform, and much less time-consuming. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|