Genetic Engineering & Biotechnology News

DEC 2018

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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GENengnews.com | Genetic Engineering & Biotechnology News | DECEMBER 2018 | 13 Douglas Horejsh, Ph.D. Senior Research Scientist, Promega Tool Provider Commentary To get a sense of how tool providers are helping researchers obtain reproducible re- sults for miRNA quantitation, GEN's editor in chief, John Sterling, posed a few questions to Douglas Horejsh, Ph.D., a senior research scientist at Promega, and Stephen Angeloni, Ph.D., a senior field applications scientist at Luminex. GEN: What types of tools and reagents are critical for miRNA research? Dr. Horejsh: Effective, safe, and reproducible miRNA purification tools are critical in the process. Obviously, it is very difficult to perform biomarker discovery if there is purification bias for particular miRNAs or inefficient dissociation of the miRNA from protein complexes. Promega has focused recent new product development efforts to purify total RNA including miRNA from tissues, plas- ma, serum, and exosomes. Many of these workflows are also automated, allowing for scaling of throughput to suit the needs of re- search laboratories. Dr. Angeloni: For in situ hybridization detection of miRNAs, proper sample preparation and probes with good signal-to-noise ratios and specificity are critical. Several compa- nies offer a variety of locked nucleic acid (LNA) probes for in situ detection of miR- NAs along with protocols and reagents for sample processing. For PCR and sequenc- ing technologies, the first limiting factor is the isolation of sufficient quantities of high- quality RNA from different samples. For some of these molecular technologies, the next critical factor is to have efficient am- plification linkers or good stem-loop prim- ers to accurately convert the miRNA into a cDNA that can be used for analysis. GEN: What are some limitations of the tools and reagents currently being used in miRNA research? Dr. Horejsh: The current standard method for purifica- tion of exosomes is ultracentrifugation. This method is reliable, but it is also tedious and time consuming. It is also not amenable to high throughput. Reagents that can be used to pre- cipitate exosomes without ultracen- trifugation would provide faster and automatable exo- some purification. However, there ap- pear to be differences in miRNA expression based on the exosome purification method. Additionally, most miRNA purification workflows currently on the market are com- pletely manual. It is very difficult to scale this type of workflow to large biomarker discov- ery or research projects. Dr. Angeloni: As more miRNA markers are associated with different diseases and become important in diagnostics, the need to detect more miRNA markers in small samples is increasing. For several PCR-based technologies, it is not easy to distinguish miRNAs that may differ by a single nucleotide. In addition, the quantity of high-quality RNA that can be isolated from various body fluids (urine, sera, other body fluids) can be too low to identify miRNAs at low concentrations accurately. When vital miRNA markers can vary by a single nucleo- tide, these factors can be a serious limitation for some diagnostic applications. HUMAN DEDICATION. MACHINE REPRODUCIBILITY. Automate your company's processes early with Terumo BCT— and we'll help take your dedication even further. ©2018 Terumo BCT, Inc. / PN 306650543A For decades, Terumo BCT has provided critical tools for committed biotech leaders like you. Today, we're bringing that partnership to new levels with our automated cell processing technologies. Our team of automation experts can help you integrate the equipment you need to pursue new standards of quality, to improve consistency and to scale your next-generation cell and gene therapies more quickly—all while helping to stay true to your budgets and timelines. And even more innovation is on the way. Let Terumo BCT help you achieve therapeutic çVWXWJSVXLITEXMIRXW[LSRIIHXLIQQSWX Learn more at XIVYQSFGXGSQQEGLMRIIéGMIRG] See miRNA Proling on page 15 OMICS ",WLVYHU\GLFXOWWR SHUIRUPELRPDUNHU GLVFRYHU\LIWKHUHLV SXULÀFDWLRQELDVIRU SDUWLFXODUPL51$Vµ —Douglas Horejsh, Ph.D.

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