Genetic Engineering & Biotechnology News

JUN15 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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18 | JUNE 15, 2018 | Genetic Engineering & Biotechnology News | Discover what's new from the leading journal on post-genomic medicine and integrative biology Sign Up for TOC Alerts Discover what's new from the leading journal on post-genomic medicine and integrative biology Sign Up for TOC Alerts Editor-in-Chief: Vural Ă–zdemir, MD, PhD, DABCP with different lineage histories within the same patient. Dark Matter "There is a good reason why we refer to the noncoding genome as dark matter," commented Gary Hon, Ph.D., assistant pro- fessor of obstetrics and gynecology at UT Southwestern Medical Center. "We know it is there. We know it plays important roles in development and disease. But we have little clue what most of it does. "Thanks to the efforts of consortia like ENCODE [the Encyclopedia of DNA Ele- ments, a project that was launched by the Na- tional Human Genome Research Institute in 2003], a map of the noncoding genome and what it encodes now exists. But despite hav- ing a great map, the single greatest challenge is that the vast majority of the millions of non- coding elements have no known function." This is a big problem that Dr. Hon's group approaches from a systems biology perspec- tive. The group invests heavily in developing new biotechnological tools to functionally in- terrogate the noncoding genome using single- cell approaches. According to Dr. Hon, the best way to understand the genome is to systematically perturb as much of it as possible. By directly measuring the functions of thousands of noncoding regions, Dr. Hon and colleagues mean to derive the fundamental principles that govern these regions. Recently developed techniques, such as Perturb-Seq, CRISPR-Seq, CROP-Seq, and Mosaic-Seq, will play important roles. The key feature they share is the ability to detect CRISPR-mediated genetic perturbations in individual cells, as well as the phenotypic consequences of these perturbations. Togeth- er with recent developments enabling thou- sands, or even millions, of single cells to be individually sequenced, these approaches are scaling the functional perturbation of the ge- nome, raising the technique to new heights. In a single well-controlled experiment, Mosaic-Seq was used to directly measure the activities of 71 enhancers. Surprisingly, even though many of these regions are predicted to be strong enhancers based on certain epigenetic features, most enhancers were found to exhibit no functional activity when repressed alone. Often, multiple enhancers had to be re- pressed to elicit a functional effect, suggesting that many enhancers are functionally redun- dant. This redundancy means that under- standing enhancer function will likely require perturbing multiple enhancers in the same cell, and further underscores the importance of applying systematic single-cell techniques like Mosaic-Seq to study these regions. Technological innovations have posi- tioned the single-cell genomics field to make great strides in the next few years, enabling a deeper understanding how the dark mat- ter of the human genome functions. Just as the ENCODE project systematically mapped millions of noncoding elements, there will be a rapid acceleration of efforts to functionally test these regions. Single-Cell Analysis Continued from page 17 OMICS Cancer is heterogeneous and understanding how multiple DNA mutations co-occur during disease pro- gression is critical to better informing therapy. Current next generation sequencing approaches lack the high sensitivity and single-cell resolution to unambiguously identify rare subclones with co-occurring mutations necessary to fully understand the complexity of the disease, according to Dennis Eastburn, Ph.D., chief sci- entific officer and co-founder, Mission Bio. Targeted single-cell DNA analysis offers the opportuni- ty to resolve genetic heterogeneity, he explains. However, methods to characterize heterogeneity and mutational status at single-cell resolution have lagged behind RNA expression analysis due to the challenges of accessing and amplifying genomic DNA. Because single-nucleotide variants (SNVs) and indels are especially informative for selecting gene specific targeted therapies, it is essential to directly assess their mutational status within a cell. "The Tapestri Platform for single-cell DNA analysis provides accurate and targeted identification of SNVs and indels to enable precise variant calling, thereby overcom- ing the incomplete genome coverage and artifactual mu- tations introduced from single-cell whole genome am- plification approaches," says Dr. Eastburn. "This targeted approach provides a robust and cost-effective solution for clinically relevant and actionable information." Work from leading cancer centers using this new precision genomics platform has found that subclones responsible for expanding and driving relapse can be found at very low levels, below 0.1 percent of the tu- mor population, prior to treatment, he continues. "Significantly, these rare subclones were undetect- able with bulk sequencing. This high-resolution view of clonal architecture has the potential to alter thera- py selection and improve disease outcomes," says Dr. Eastburn. n High-Throughput Targeted Single-Cell DNA Analysis

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