effective method of DNA extraction for bioleaching bacteria from acid mine drainage

An effective and versatile method for microorganism lysis and direct extraction of DNA from bioleaching bacteria was developed using pure cultures and an acid mine drainage (AMD) sediment sample. In the described method, microorganisms are treated at three different incubation temperatures: boiling...

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Bibliographic Details
Published in:Applied microbiology and biotechnology 2008-07, Vol.79 (5), p.881-888, Article 881
Main Authors: Zeng, Leping, Huang, Jufang, Zhang, Yanfei, Qiu, Guanzhou, Tong, Jianbin, Chen, Dan, Zhou, Jin, Luo, Xuegang
Format: Article
Language:English
Subjects:
Acid Mine Drainage
Acidithiobacillus - genetics
Acidithiobacillus - isolation & purification
Alcohol
Bacteria
Bacterial Proteins - genetics
Biohydrometallurgy. Microbial leaching
Bioleaching bacteria
Biological and medical sciences
Biotechnology
DNA extraction
DNA Fingerprinting
DNA Gyrase - genetics
DNA, Bacterial - genetics
DNA, Bacterial - isolation & purification
DNA, Ribosomal - genetics
DNA, Ribosomal - isolation & purification
Fundamental and applied biological sciences. Psychology
Gene amplification
Genetic Techniques - economics
Geologic Sediments - microbiology
Industrial applications and implications. Economical aspects
Leaching
Life Sciences
Methods
Microbial Genetics and Genomics
Microbiology
Microorganisms
Mining
Molecular Sequence Data
PCR-RFLP
Phenols
Polymerase Chain Reaction
Polymorphism
Rep-PCR
RNA, Ribosomal, 16S - genetics
RNA, Ribosomal, 16S - isolation & purification
Sediments
Studies
Water Microbiology
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Summary:An effective and versatile method for microorganism lysis and direct extraction of DNA from bioleaching bacteria was developed using pure cultures and an acid mine drainage (AMD) sediment sample. In the described method, microorganisms are treated at three different incubation temperatures: boiling water incubation for 6-10min, followed by 60 ± 5°C for 30min, then 72°C for 30min. The extracted DNA is purified using a phenol/chloroform/alcohol mixture and precipitated in absolute alcohol. The 16S ribosomal RNA (rRNA) and gyrB genes of the pure cultures were amplified using the polymerase chain reaction (PCR) and differentiated using repetitive intergenic DNA sequences amplification (Rep-PCR). For the AMD sediment sample, the 16S rRNA and gyrB genes of the amplicons were digested with Hin6I and MspI, and the restriction fragment length polymorphism analysis patterns were used as a fingerprint to discern community diversity. The results indicated that this method is a versatile, reproducible, effective, and rapid technique for routine DNA extraction from bioleaching bacteria. The low cost of this method also makes it attractive for large-scale studies.
ISSN:0175-7598
1432-0614
DOI:10.1007/s00253-008-1491-5