Genetic Engineering & Biotechnology News

NOV15 2017

Genetic Engineering & Biotechnology News (GEN) is the world's most widely read biotech publication. It provides the R&D community with critical information on the tools, technologies, and trends that drive the biotech industry.

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Genetic Engineering & Biotechnology News | | NOVEMBER 15, 2017 | 23 bringing STR technology to another species," said Maryellen de Mars, Ph.D., vice president, Standards Resource Center, ATCC. In 2012, ATCC's Standards Development Organization released a consensus standard for authenticating human cell lines using STR analysis. In addition to identifying amelogenin and 8 autosomal STR loci as markers for au- thentication, the standard also recommends a minimum matching criterion of 80% to de- termine if two cell lines are really the same. According to Dr. de Mars, it will "take time and effort" to develop a similar standard for mouse cell line authentication: "[MCLAC] needs to identify the correct combination of loci needed to sufficiently distinguish mouse cell lines using STR analysis." Since offspring can inherit specific STR polymorphisms from their parents, the sub- stantial inbreeding of mice by the scientific community means that researchers will need more markers to distinguish between cell lines: thus far MCLAC has established a com- bination of 19 mouse STR loci and 2 human markers. In addition to identifying robust STR markers, MCLAC will also need to generate a mouse cell line database, establish an algo- rithm for comparing STR profiles, and define matching criterion. In the meantime, ATCC offers a mitochon- drial cytochrome c oxidase subunit 1 (CO1) assay that uses DNA barcoding to discrimi- nate between species, which researchers can use both to confirm their cell lines at the spe- cies level and to detect interspecies contami- nation (Figure 3). "Authentication actually involves both identity and purity," explained Dr. de Mars. In addition to identifying the cell line, authentication should test for cross-con- tamination between cell lines, as well as viral and mycoplasma contamination. Catching the Genetic Drift (Evolution in a Petri Dish) Although cell culturists typically think of contamination in terms of microorgan- isms—bacteria, fungi, viruses, and proto- zoa—cross-contamination between cell lines poses a particularly worrisome threat. Con- tamination events that introduce stray cells from another culture can go unnoticed. And unnoticed problems can multiply. As Steve Jackson, Ph.D., application sci- entist at Thermo Fisher Scientific remarked, "If something has a growth advantage, even a small contaminant is going to overgrow the cell line in a very short time." Nothing exemplifies this truth more than the HeLa cell line, which the International Cell Line Authentication Committee (ICLAC) has identified as a con- taminant in 113 different cell lines (Figure 3). Not only can natural selection lead to a hostile takeover of your culture by HeLa cells, but it can also lead to genetic drift within the population. "We're talking about evolution in a Petri dish," Dr. Jackson told GEN, and spontaneous mutations can give rise to sub- populations of genetic variants within the cell line. Immortalized and tumor cell lines may have an even greater proclivity toward genetic changes, because they often go through chro- mosomal rearrangements. To monitor cell lines for genetic drift, Da- vid Yoder, Ph.D., a field application scientist at Thermo Fisher Scientific, recommends that scientists use STR assays that interro- gate a broad range of STR markers including Thermo Fisher Scientific's AuthentiFiler™, Indentifiler™ Plus, Indentifiler™ Direct, and GlobalFiler™. In addition to amplifying am- elogenin, these PCR amplification kits con- tain primers for 9, 15, 15, and 21 different autosomal STR markers, respectively. "Having a large panel doesn't guarantee you will detect these [genetic] changes," ad- mitted Dr. Yoder, "but it does increase the opportunity to detect them." Both Gen- eMapper ® ID-X and GeneMapper ® 5 Soft- ware, available through Thermo Fisher Sci- entific, allow users to process and compare STR data between samples to identify and WE BRING LIFE TO YOUR LABORATORY. Explore & get your free demo at Shakers | Bioreactors | Bioprocess Platform Software Connect and manage different systems. One single solution to unify all your equipment and data. eve ® – the revolutionary bioprocess platform software. © Space Telescope Science Institute See Cell Line Authentication on page 24 Bioprocessing Figure 3. The aggressive HeLa cell line, shown in this fluorescent microscopy image with cytoskeletal (magenta) and nuclear (cyan) components highlighted, looks similar to other cell lines, but it's one of the most common contaminants identified in ICLAC's Register of Misidentified Cell Lines. To confirm the identity of its cell lines, ATCC has used DNA polymorphisms in addition to enzyme polymorphisms, HLA typing, and karyotyping. Such work may require STR profiles, which are available for all ATCC human cell lines.

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