Rectal swab DNA collection protocol for PCR genotyping in rats
Creators: name: Kaye, Audrey E
name: Proctor-Bonbright, Jacob W
name: Yu, Jai Y
Publisher: name: Taylor & Francis
Year: 2024
Subjects: subject: Medicine
subject: Microbiology
subject: FOS: Biological sciences
subject: Cell Biology
subject: Genetics
subject: Molecular Biology
subject: Pharmacology
subject: Biotechnology
subject: Chemical Sciences not elsewhere classified
subject: Cancer
subject: Science Policy
subject: Infectious Diseases
subject: FOS: Health sciences
subject: Virology

description: DNA collection is essential for genotyping laboratory animals. Common collection methods require tissue amputation, causing discomfort and injury. Rectal swabbing has been proposed as an effective, minimally invasive alternative, but an evidence-backed protocol for the technique remains unavailable. This report evaluates the effects of collection parameters on the quality of PCR results and presents a protocol for genotyping a litter of rats within 3–5 h. Samples with 2–8 scrapes produced enough DNA to amplify targets up to ∼1800 bp long using PCR. Rectal swabbing produced PCR results with similar utility as ear clip samples, and results were unaffected by residual fecal matter or cell debris. The protocol enables fast, minimally invasive and repeatable genotyping using commercial PCR reagents. DNA collection for genotyping laboratory rodents commonly requires tissue amputation, leading to injury and discomfort. Rectal swabbing can be an effective, minimally invasive alternative, but there is a lack of information on how to apply the technique and the necessary parameters involved. This report assesses collection and processing parameters while combining the rectal swab collection method with commercial PCR reagents to allow for fast genotyping with minimally processed DNA samples. Findings were used to design a protocol for minimally invasive DNA collection and genotyping. Rectal swabbing is an effective yet minimally invasive technique for collecting DNA from laboratory rats. This new protocol consistently produces PCR-quality DNA while remaining repeatable and cost effective. #Genotyping #PCR #Transgenic Various parameters of the rectal swabbing technique for DNA collection were evaluated. These parameters included the number of rectal scrapes, number of samples, and contamination with fecal matter or cell debris. Samples with 2–8 scrapes yielded sufficient DNA for PCR amplification of genomic sequences up to ∼1800 bp. Contamination with fecal matter or cell debris did not affect PCR outcome quality. Rectal swabbing produced similar results across rats of both sexes and different ages. Common DNA collection techniques for genotyping laboratory rodents require tissue amputation, causing injury and discomfort. Rectal swabbing can be an effective, minimally invasive alternative but is not commonly used. There is no existing protocol for performing the procedure. Rectal swab collection was conducted with genetically modified male and female rats of various ages. The number of samples, number of rectal scrapes per sample and presence of fecal matter or cell debris were experimentally varied to determine their effect on PCR amplification. A total of 2–8 scrapes of the rectal epithelium can yield sufficient DNA for the PCR amplification of genomic targets up to ∼1800 bp. Fecal matter and cell debris do not affect PCR outcome quality. A detailed protocol for this procedure is provided. Rectal swabbing is an effective and minimally invasive alternative for DNA collection in rats.