A Rapid and Reproducible Genomic DNA Extraction Protocol for Sequence-Based Identification of Archaea, Bacteria, Cyanobacteria, Diatoms, Fungi, and Green Algae

  • Farkhondeh Saba Microorganisms Bank, Iranian Biological Resource Center (IBRC), (ACECR), Tehran, Iran.
  • Moslem Papizadeh Microorganisms Bank, Iranian Biological Resource Center (IBRC), (ACECR), Tehran, Iran.
  • Javad Khansha Microorganisms Bank, Iranian Biological Resource Center (IBRC), (ACECR), Tehran, Iran.
  • Mahshid Sedghi Microorganisms Bank, Iranian Biological Resource Center (IBRC), (ACECR), Tehran, Iran.
  • Mehrnoosh Rasooli Microorganisms Bank, Iranian Biological Resource Center (IBRC), (ACECR), Tehran, Iran.
  • Mohammad Ali Amoozegar 2 Extremophiles Laboratory, Department of Microbiology, Faculty of Biology and Center of Excellence in Phylogeny of Living Organisms, College of Science, University of Tehran, Tehran, Iran.
  • Mohammad Reza Soudi National Laboratory of Industrial Microbiology, Department of Microbiology, Faculty of Biological Sciences, Alzahra University, Tehran, Iran.
  • Seyed Abolhassan Shahzadeh Fazeli Department of Molecular and Cellular Biology, Faculty of Basic Sciences and Advanced Technologies in biology, University of Science and Culture, Tehran, Iran.
Keywords: Eukaryotes, Genome (DNA) extraction, Identification, Prokaryotes, Sequence

Abstract

Background:  Sequence-based identification of various microorganisms including Archaea, Bacteria, Cyanobacteria, Diatoms, Fungi, and green algae necessitates an efficient and reproducible genome extraction procedure though which a pure template DNA is yielded and it can be used in polymerase chain reactions (PCR). Considering the fact that DNA extraction from these microorganisms is time consuming and laborious, we developed and standardized a safe, rapid and inexpensive miniprep protocol. Methods:  According to our results, amplification of various genomic regions including SSU, LSU, ITS, β-tubulin, actin, RPB2, and EF-1 resulted in a reproducible and efficient DNA extraction from a wide range of microorganisms yielding adequate pure genomic material for reproducible PCR-amplifications. Results:   This method relies on a temporary shock of increased concentrations of detergent which can be applied concomitant with multiple freeze-thaws to yield sufficient amount of DNA for PCR amplification of multiple or single fragments(s) of the genome. As an advantage, the recipe seems very flexible, thus, various optional steps can be included depending on the samples used.Conclusion:   Having the needed flexibility in each step, this protocol is applicable on a very wide range of samples. Hence, various steps can be included depending on the desired quantity and quality.

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Published
2017-01-13
How to Cite
1.
Saba F, Papizadeh M, Khansha J, Sedghi M, Rasooli M, Amoozegar MA, Soudi MR, Shahzadeh Fazeli SA. A Rapid and Reproducible Genomic DNA Extraction Protocol for Sequence-Based Identification of Archaea, Bacteria, Cyanobacteria, Diatoms, Fungi, and Green Algae. J Med Bacteriol. 5(3-4):22-8.
Section
Original Articles