Comparison of DNA Extraction Efficiency and Reproducibility of Different Aeration Diffuser Biofilms Using Bead-Beating Protocol

Pitiporn Asvapathanagul, Manel Garrido-Baserba, Betty H. Olson, Hee-Deung Park, Deqiang Chen, Diego Rosso

Research output: Contribution to journalArticle

Abstract

An existing bead-beating DNA extraction protocol was employed to compare the DNA extraction recovery and fragment quality of 6 different aeration diffuser biofilms. Escherichia coli, Gordonia amarae, and mixed liquor were used as controls. The fraction of total DNAbiofilm decreased monotonically with increasing number of beat beatings (BB) when the amount of DNA present was sufficient (>4 μgDNA/cm2), excluding the ceramic disk. While controls required only 2 BBs, 3 out of 5 BBs achieved ≥70% of total DNA (70.3 ± 1.7%) for 5 out of 6 biofilms. Quantitative polymerase chain reaction (PCR) analyses of 353 and 1,505 basepair (bp) amplicons from pure culture extracts showed target copy numbers were not degraded for the first 2 BBs, but the third BB decreased amplicon concentrations by 0.65 and 1.12 log for E. coli, and 0.39 and 0.40 log for G. amarae, respectively. The 353 bp fragment amplification from biofilm samples showed minimal degradation for the first 3 BBs. PCR and gel electrophoresis confirmed integrity of amplified 1,505 bp DNA fragments over the 5 BBs, except in the EDPM (75 mm diameter, tube) diffuser biofilm (4.98 ± 0.62 μgDNA/cm2). Taken together, this study showed type of diffuser membrane biofilms had no effects on extraction efficiency, but low DNA concentrations reduced extraction performance.

Original languageEnglish
Pages (from-to)293-303
Number of pages11
JournalJournal of Molecular Microbiology and Biotechnology
Volume28
Issue number6
DOIs
Publication statusPublished - 2019 Jul 1

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Keywords

  • Aeration
  • Bead beating
  • Biofilm
  • DNA extraction efficiency
  • Fine-pore diffusers
  • Fragment length amplification
  • Wastewater

ASJC Scopus subject areas

  • Biotechnology
  • Microbiology
  • Applied Microbiology and Biotechnology
  • Molecular Biology

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