Skip to main content
ARS Home » Plains Area » Lincoln, Nebraska » Agroecosystem Management Research » Research » Publications at this Location » Publication #424469
Research Project: Managing Manure as a Soil Resource for Improved Biosecurity, Nutrient Availability, and Soil Sustainability

Location: Agroecosystem Management Research

Title: 4-Gene standard uniplex PCR SOP for target antibiotic resistance genes

Author
item Condon, Justine
item Durso, Lisa

Submitted to: Protocols.io
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/1/2025
Publication Date: 4/29/2025
Citation: Condon, J.C., Durso, L.M. 2025. 4-Gene standard uniplex PCR SOP for target antibiotic resistance genes. Protocols.io. https://doi.org/10.17504/protocols.io.n92ldrxnng5b/v1.
DOI: https://doi.org/10.17504/protocols.io.n92ldrxnng5b/v1

Interpretive Summary: When people are sick with infectious diseases, antibiotics are commonly used to treat and cure the infection.  However more and more infectious disease-causing bacteria are becoming resistant to antibiotics, resulting in people being sick longer, or dying when they once might have survived.  Because of this, antibiotic resistance is a global health crisis.  This protocol describes an assay that probes the bacterial DNA to identify if they carry the instructions needed for antibiotic resistance, using a technique called Polymerase Chain Reaction (PCR).

Technical Abstract: Purpose This protocol describes the correct procedures on how to set-up polymerase chain reaction (PCR) and run gel electrophoresis for selected antibiotic resistance genes and determinants: sulfonamide (sul), erythromycin (erm(B)), extended spectrum ß-lactamase (blaCTX-M-1), integrase (intI1), and tetracycline tet(A), tet(O), and tet(X). Scope Polymerase Chain Reaction (PCR) is a technique that enables researchers to rapidly and efficiently amplify (copy) a specific region of DNA (Deoxyribonucleic Acid) or RNA (Ribonucleic Acid) into millions of copies. A simple PCR reaction consists of target DNA, a set of synthetic oligonucleotide primers that flank the target DNA sequence, a thermostable DNA polymerase (usually Taq polymerase), and nucleotides. Thermal cycling exposes these contents to repeated cycles of heating and cooling that promotes DNA replication. There are three steps involved in the process: denaturation, annealing, and elongation.