TY - JOUR
T1 - AsChip
T2 - A High-Throughput qPCR Chip for Comprehensive Profiling of Genes Linked to Microbial Cycling of Arsenic
AU - Zhao, Yi
AU - Su, Jian Qiang
AU - Ye, Jun
AU - Rensing, Christopher
AU - Tardif, Stacie
AU - Zhu, Yong Guan
AU - Brandt, Kristian Koefoed
PY - 2019/1/15
Y1 - 2019/1/15
N2 -
Arsenic (As) is a ubiquitous toxic element adversely affecting human health. Microbe-mediated cycling of As is largely mediated by detoxification and energy metabolism in microorganisms. We here report the development of a novel high-throughput qPCR (HT-qPCR) chip (AsChip) for comprehensive profiling of genes involved in microbial As cycling (here collectively termed "As genes"). AsChip contained 81 primer sets targeting 19 As genes and the 16S rRNA gene as a reference gene. Gene amplicon sequencing showed high identity (>96%) of newly designed primers corresponding to their targets. AsChip displayed high sensitivity (plasmid template serial dilution test; r = -0.99), with more than 96% of all PCR assays yielding true positive signals. R
2
coefficients for standard curves and PCR amplification efficiencies averaged 0.98 and 0.99, respectively. A high correlation between C
T
values obtained by AsChip and conventional qPCR was obtained (r = 0.962, P < 0.001). Finally, we successfully applied AsChip on soil samples from a chromium-copper-arsenic-contaminated field site and identified diverse As genes with total abundance average of 0.4 As gene copies per 16S rRNA. Our results indicate that AsChip constitutes a robust tool for comprehensive quantitative profiling of As genes in environmental samples.
AB -
Arsenic (As) is a ubiquitous toxic element adversely affecting human health. Microbe-mediated cycling of As is largely mediated by detoxification and energy metabolism in microorganisms. We here report the development of a novel high-throughput qPCR (HT-qPCR) chip (AsChip) for comprehensive profiling of genes involved in microbial As cycling (here collectively termed "As genes"). AsChip contained 81 primer sets targeting 19 As genes and the 16S rRNA gene as a reference gene. Gene amplicon sequencing showed high identity (>96%) of newly designed primers corresponding to their targets. AsChip displayed high sensitivity (plasmid template serial dilution test; r = -0.99), with more than 96% of all PCR assays yielding true positive signals. R
2
coefficients for standard curves and PCR amplification efficiencies averaged 0.98 and 0.99, respectively. A high correlation between C
T
values obtained by AsChip and conventional qPCR was obtained (r = 0.962, P < 0.001). Finally, we successfully applied AsChip on soil samples from a chromium-copper-arsenic-contaminated field site and identified diverse As genes with total abundance average of 0.4 As gene copies per 16S rRNA. Our results indicate that AsChip constitutes a robust tool for comprehensive quantitative profiling of As genes in environmental samples.
UR - http://www.scopus.com/inward/record.url?scp=85060111870&partnerID=8YFLogxK
U2 - 10.1021/acs.est.8b03798
DO - 10.1021/acs.est.8b03798
M3 - Journal article
C2 - 30532956
AN - SCOPUS:85060111870
SN - 0013-936X
VL - 53
SP - 798
EP - 807
JO - Environmental Science & Technology
JF - Environmental Science & Technology
IS - 2
ER -
