Species identification in cell culture: a two-pronged molecular approach.
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.
Cooper
Jason K
JK
ATCC (American Type Culture Collection), 10801 University Boulevard, Manassas, VA 20110, USA.
Sykes
Greg
G
King
Steve
S
Cottrill
Karin
K
Ivanova
Natalia V
NV
Hanner
Robert
R
Ikonomi
Pranvera
P
D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D003576:Electron Transport Complex IV / Q000378:metabolism; D006801:Humans; D020014:K562 Cells; D010802:Phylogeny; D021122:Protein Subunits / Q000378:metabolism; D013045:Species Specificity
D021122:Protein Subunits; D003576:Electron Transport Complex IV