TY - JOUR
T1 - Comparison of DNA Extraction Efficiency and Reproducibility of Different Aeration Diffuser Biofilms Using Bead-Beating Protocol
AU - Asvapathanagul, Pitiporn
AU - Garrido-Baserba, Manel
AU - Olson, Betty H.
AU - Park, Hee Deung
AU - Chen, Deqiang
AU - Rosso, Diego
N1 - Funding Information:
This study was funded by Water Environment Research Foundation (WERF Project Number: INFR2R12) and the United States Environmental Protection Agency (EPA-WERF Cooperative Agreement No. CR-83419201). Additionally, the Mentor Appreciation Fund provided by the National Institute of General Medical Sciences of the National Institutes of Health under Award Numbers; UL1GM118979 encouraged the manuscript publication.
Funding Information:
This study was funded by Water Environment Research Foundation (WERF, Project Number: INFR2R12) and the United States Environmental Protection Agency (EPA-WERF Cooperative Agreement No. CR-83419201), with the support of the Irvine Ranch Water District, DC Water, Hampton Roads Sanitation District, Sydney Water, and Southern California Edison. The authors thank David M. Hayden of the Irvine Ranch Water District at the Michelson Water Reclamation Plant for supporting the reactor installation and the sampling campaign and Alice K. Robinson of BKT and Graham W. McCarthy of the Civil and Environmental Engineering Department, University of California, Irvine for their assistant during sampling.
Publisher Copyright:
© 2019 S. Karger AG, Basel. All rights reserved.
PY - 2019/7/1
Y1 - 2019/7/1
N2 - 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.
AB - 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.
KW - Aeration
KW - Bead beating
KW - Biofilm
KW - DNA extraction efficiency
KW - Fine-pore diffusers
KW - Fragment length amplification
KW - Wastewater
UR - http://www.scopus.com/inward/record.url?scp=85068623260&partnerID=8YFLogxK
U2 - 10.1159/000500875
DO - 10.1159/000500875
M3 - Article
C2 - 31234170
AN - SCOPUS:85068623260
SN - 1464-1801
VL - 28
SP - 293
EP - 303
JO - Journal of Molecular Microbiology and Biotechnology
JF - Journal of Molecular Microbiology and Biotechnology
IS - 6
ER -