Species identification in cell culture: a two-pronged molecular approach |
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Authors: | Jason K Cooper Greg Sykes Steve King Karin Cottrill Natalia V Ivanova Robert Hanner Pranvera Ikonomi |
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Institution: | (1) ATCC (American Type Culture Collection), 10801 University Boulevard, Manassas, VA 20110, USA;(2) Biodiversity Institute of Ontario, University of Guelph, Guelph, ON, Canada, N1G 2W1 |
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Abstract: | Species identification of cell lines and detection of cross-contamination are crucial for scientific research accuracy and
reproducibility. Whereas short tandem repeat profiling offers a solution for a limited number of species, primarily human
and mouse, the standard method for species identification of cell lines is enzyme polymorphism. Isoezymology, however, has
its own drawbacks; it is cumbersome and the data interpretation is often difficult. Furthermore, the detection sensitivity
for cross-contamination is low; it requires large amounts of the contaminant present and cross-contamination within closely
related species may go undetected. In this paper, we describe a two-pronged molecular approach that addresses these issues
by targeting the mitochondrial genome. First, we developed a multiplex PCR-based assay to rapidly identify the most common
cell culture species and quickly detect cross-contaminations among these species. Second, for speciation and identification
of a wider variety of cell lines, we amplified and sequenced a 648-bp region, often described as the “barcode region” by using
a universal primer mix targeted at conserved sequences of the cytochrome C oxidase I gene (COI). This method was challenged
with a panel of 67 cell lines from 45 diverse species. Implementation of these assays will accurately determine the species
of cell lines and will reduce the problems of misidentification and cross-contamination that plague research efforts. |
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Keywords: | DNA barcode Sequencing PCR Cytochrome C oxidase I (COI) Species-specific primers |
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