Rapid dissection and model-based optimization of inducible enhancers in human cells using a massively parallel reporter assay

Learning to read and write the transcriptional regulatory code is of central importance to progress in genetic analysis and engineering. Here we describe a massively parallel reporter assay (MPRA) that facilitates the systematic dissection of transcriptional regulatory elements. In MPRA, microarray-synthesized DNA regulatory elements and unique sequence tags are cloned into plasmids to generate a library of reporter constructs. These constructs are transfected into cells and tag expression is assayed by high-throughput sequencing. We apply MPRA to compare >27,000 variants of two inducible enhancers in human cells: a synthetic cAMP-regulated enhancer and the virus-inducible interferon-β enhancer. We first show that the resulting data define accurate maps of functional transcription factor binding sites in both enhancers at single-nucleotide resolution. We then use the data to train quantitative sequence-activity models (QSAMs) of the two enhancers. We show that QSAMs from two cellular states can be combined to design enhancer variants that optimize potentially conflicting objectives, such as maximizing induced activity while minimizing basal activity.National Human Genome Research Institute (U.S.) (grant R01HG004037)National Science Foundation (U.S.) ((NSF) grant PHY-0957573)National Science Foundation (U.S.) (NSF grant PHY-1022140)Broad Institut

QCD and strongly coupled gauge theories : challenges and perspectives

We highlight the progress, current status, and open challenges of QCD-driven physics, in theory and in experiment. We discuss how the strong interaction is intimately connected to a broad sweep of physical problems, in settings ranging from astrophysics and cosmology to strongly coupled, complex systems in particle and condensed-matter physics, as well as to searches for physics beyond the Standard Model. We also discuss how success in describing the strong interaction impacts other fields, and, in turn, how such subjects can impact studies of the strong interaction. In the course of the work we offer a perspective on the many research streams which flow into and out of QCD, as well as a vision for future developments.Peer reviewe

Effective preparation of Plasmodium vivax field isolates for high-throughput whole genome sequencing.

Whole genome sequencing (WGS) of Plasmodium vivax is problematic due to the reliance on clinical isolates which are generally low in parasitaemia and sample volume. Furthermore, clinical isolates contain a significant contaminating background of host DNA which confounds efforts to map short read sequence of the target P. vivax DNA. Here, we discuss a methodology to significantly improve the success of P. vivax WGS on natural (non-adapted) patient isolates. Using 37 patient isolates from Indonesia, Thailand, and travellers, we assessed the application of CF11-based white blood cell filtration alone and in combination with short term ex vivo schizont maturation. Although CF11 filtration reduced human DNA contamination in 8 Indonesian isolates tested, additional short-term culture increased the P. vivax DNA yield from a median of 0.15 to 6.2 ng µl(-1) packed red blood cells (pRBCs) (p = 0.001) and reduced the human DNA percentage from a median of 33.9% to 6.22% (p = 0.008). Furthermore, post-CF11 and culture samples from Thailand gave a median P. vivax DNA yield of 2.34 ng µl(-1) pRBCs, and 2.65% human DNA. In 22 P. vivax patient isolates prepared with the 2-step method, we demonstrate high depth (median 654X coverage) and breadth (≥89%) of coverage on the Illumina GAII and HiSeq platforms. In contrast to the A+T-rich P. falciparum genome, negligible bias was observed in coverage depth between coding and non-coding regions of the P. vivax genome. This uniform coverage will greatly facilitate the detection of SNPs and copy number variants across the genome, enabling unbiased exploration of the natural diversity in P. vivax populations

Climate change and zoonotic infections in the Russian Arctic

Climate change in the Russian Arctic is more pronounced than in any other part of the country. Between 1955 and 2000, the annual average air temperature in the Russian North increased by 1.2°C. During the same period, the mean temperature of upper layer of permafrost increased by 3°C. Climate change in Russian Arctic increases the risks of the emergence of zoonotic infectious diseases. This review presents data on morbidity rates among people, domestic animals and wildlife in the Russian Arctic, focusing on the potential climate related emergence of such diseases as tick-borne encephalitis, tularemia, brucellosis, leptospirosis, rabies, and anthrax