Mouse cell line authentication

Abstract

The scientific community has responded to the misidentification of human cell lines with validated methods to authenticate these cells; however, few assays are available for nonhuman cell line identification. We have developed a multiplex polymerase chain reaction assay that targets nine tetranucleotide short tandem repeat (STR) markers in the mouse genome. Unique profiles were obtained from seventy-two mouse samples that were used to determine the allele distribution for each STR marker. Correlations between allele fragment length and repeat number were determined with DNA Sanger sequencing. Genotypes for L929 and NIH3T3 cell lines were shown to be stable with increasing passage numbers as there were no significant differences in fragment length with samples of low passage when compared to high passage samples. In order to detect cell line contaminants, primers for two human STR markers were incorporated into the multiplex assay to facilitate detection of human and African green monkey DNA. This multiplex assay is the first of its kind to provide a unique STR profile for each individual mouse sample and can be used to authenticate mouse cell lines.

Molecular characterization of mouse lens epithelial cell lines and their suitability to study RNA granules and cataract associated genes

“…Richard Maas and Gary Laverty, respectively. To further determine the authenticity of the cell lines as recommended by the eye research community, we used mouse and human-specific primer sets designed to detect STRs within the respective genomes as previously described (Almeida et al, 2014). As expected, all four cell lines exhibit amplification of mouse STR region-specific PCR products, while no PCR product with the human genome-specific STR primer sets is detected (Figure 1).…”

“…Primers were chosen for authentication based on murine and human short tandem repeats (STRs) within their respective genomes as recommended (Almeida et al, 2014). The two human primers D4S2408 and D8S1106 are abbreviated to HD4S and HD8S respectively within this publication.…”

“…To validate their species identity, PCR on genomic DNA from individual cell lines was used to test the presence or absence of mouse or human-specific short tandem repeats (STRs) as previously recommended (Almeida et al, 2014). Mouse-specific PCR products are indicated by their chromosomal location (X-1 through 18-3).…”

Microbial and Viral Contamination of Animal and Stem Cell Cultures: Common Contaminants, Detection and Elimination

“…A series of STR markers that distinguish African green monkey, Chinese hamster and mouse cell lines have been identified. 28 Nine of those markers have been successfully used in multiplex PCR assay. 29 The isoenzyme analysis, 30 karyotyping 31 and DNA barcoding (mitochondrial DNA typing) 32 are some other commonly used methods to identify and verify the species of origin.…”

Cell line authentication to improve preclinical cancer research: Methods in cell line authentication, quality control, and annotation

“…Although mouse STR primers are also available, a standardized STR test for murine has yet to be developed. A challenge remains that many murine cell lines are derived from a handful of inbred strains; however, a recent SNP-based analysis indicates a test could be developed (19,20). Therefore, the chance of detecting mouse intra-species cross-contamination is low and development of a reliable test is needed.…”