| Abstract |
Species identification is an integral part of cell culture authentication
and helps identify cross-contaminated or misidentified animal cell lines.
Karyotyping is time-consuming, expensive, and may require data analysis
expertise. Isoenzyme analysis, the gold standard for animal species
identification, is no longer an option with discontinued reagent
production. Here, two species identification consensus standards are
recommended to replace isoenzymology. In the polymerase chain reaction
(PCR)-based assay, primers target mitochondrial genome regions of fifteen
of the most common species employed in cell culture. A sample
identification PCR confirms the putative species; a cross contamination
multiplex uses the full primer set except for the primers targeting the
listed culture species and amplifies any contaminant DNA present. Animal
cells from fish, insects, and other species not covered in the PCR assay
may be identified through sequence-based "DNA Barcoding," which examines a
648-bp region of the semi-conserved cytochrome oxidase I (CO1) gene after
amplification via universal primers. The resulting sequence data are
compared against a database, populated with high quality-score reference
sequences, to identify the species. The previously published DNA Barcode
(ASN-0003) and the PCR-Based (ASN-0004) consensus standards under
development are two complementary analytical techniques facilitating
species identification of anything from research to production-scale cell-
based applications.
|
|---|
