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Identification and quantification of arsC genes in environmental samples by using real-time PCR

TitleIdentification and quantification of arsC genes in environmental samples by using real-time PCR
Publication TypeJournal Article
Year of Publication2004
AuthorsSun, YM, Polishchuk, EA, Radoja, U, Cullen, WR
JournalJournal of Microbiological Methods
Volume58
Pagination335-349
Date PublishedSep
Type of ArticleArticle
ISBN Number0167-7012
KeywordsarsC gene, arsenate reduction, ASSAY, BACTERIA, DNA, ESCHERICHIA-COLI, group-specific primers, QUANTITATION, real-time PCR, REDUCTION, resistance, SOIL, SYBR (R) Green 1
Abstract

The arsC gene is responsible for the first step in arsenate biotransformation encoding the enzyme arsenate reductase. The quantitative real-time PCR method was developed to quantify the abundance of the arsC genes in environmental samples contaminated with arsenic. Two sets of primers that showed high specificity for the target arsC gene were designed based on consensus sequences from 13 bacterial species. The arsC gene was used as an external standard instead of total DNA in the calibration curve for real-time PCR, which was linear over six orders of magnitude and the detection limit was estimated to be about three copies of the gene. Soil samples from arsenic contaminated sites were screened for arsC genes by using PCR and showed the presence of this gene. The copy numbers of the gene ranging from 0.88 x 10(4) to 1.56 x 10(5) per ng total DNA were found in eight arsenic contaminated samples. Soil samples from a bioreactor containing pulp mill biomass and high concentration of arsenate showed a tenfold higher count of arsC gene copies than soil samples collected underground from an arsenic-rich gold mine. (C) 2004 Elsevier B.V. All rights reserved.

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