CrY2H-seq: a massively multiplexed assay for deep-coverage interactome mapping.

Broad-scale protein-protein interaction mapping is a major challenge given the cost, time, and sensitivity constraints of existing technologies. Here, we present a massively multiplexed yeast two-hybrid method, CrY2H-seq, which uses a Cre recombinase interaction reporter to intracellularly fuse the coding sequences of two interacting proteins and next-generation DNA sequencing to identify these interactions en masse. We applied CrY2H-seq to investigate sparsely annotated Arabidopsis thaliana transcription factors interactions. By performing ten independent screens testing a total of 36 million binary interaction combinations, and uncovering a network of 8,577 interactions among 1,453 transcription factors, we demonstrate CrY2H-seq's improved screening capacity, efficiency, and sensitivity over those of existing technologies. The deep-coverage network resource we call AtTFIN-1 recapitulates one-third of previously reported interactions derived from diverse methods, expands the number of known plant transcription factor interactions by three-fold, and reveals previously unknown family-specific interaction module associations with plant reproductive development, root architecture, and circadian coordination

Synthesis and cell-free cloning of DNA libraries using programmable microfluidics

Microfluidics may revolutionize our ability to write synthetic DNA by addressing several fundamental limitations associated with generating novel genetic constructs. Here we report the first de novo synthesis and cell-free cloning of custom DNA libraries in sub-microliter reaction droplets using programmable digital microfluidics. Specifically, we developed Programmable Order Polymerization (POP), Microfluidic Combinatorial Assembly of DNA (M-CAD) and Microfluidic In-vitro Cloning (MIC) and applied them to de novo synthesis, combinatorial assembly and cellfree cloning of genes, respectively. Proof-of-concept for these methods was demonstrated by programming an autonomous microfluidic system to construct and clone libraries of yeast ribosome binding sites and bacterial Azurine, which were then retrieved in individual droplets and validated. The ability to rapidly and robustly generate designer DNA molecules in an autonomous manner should have wide application in biological research and development

Next generation sequencing in cancer: opportunities and challenges for precision cancer medicine

Over the past decade, testing the genes of patients and their specific cancer types has become standardized practice in medical oncology since somatic mutations, changes in gene expression and epigenetic modifications are all hallmarks of cancer. However, while cancer genetic assessment has been limited to single biomarkers to guide the use of therapies, improvements in nucleic acid sequencing technologies and implementation of different genome analysis tools have enabled clinicians to detect these genomic alterations and identify functional and disease-associated genomic variants. Next-generation sequencing (NGS) technologies have provided clues about therapeutic targets and genomic markers for novel clinical applications when standard therapy has failed. While Sanger sequencing, an accurate and sensitive approach, allows for the identification of potential novel variants, it is however limited by the single amplicon being interrogated. Similarly, quantitative and qualitative profiling of gene expression changes also represents a challenge for the cancer field. Both RT-PCR and microarrays are efficient approaches, but are limited to the genes present on the array or being assayed. This leaves vast swaths of the transcriptome, including non-coding RNAs and other features, unexplored. With the advent of the ability to collect and analyze genomic sequence data in a timely fashion and at an ever-decreasing cost, many of these limitations have been overcome and are being incorporated into cancer research and diagnostics giving patients and clinicians new hope for targeted and personalized treatment. Below we highlight the various applications of next-generation sequencing in precision cancer medicine

Asymmetric Lee Distance Codes for DNA-Based Storage

We consider a new family of codes, termed asymmetric Lee distance codes, that arise in the design and implementation of DNA-based storage systems and systems with parallel string transmission protocols. The codewords are defined over a quaternary alphabet, although the results carry over to other alphabet sizes; furthermore, symbol confusability is dictated by their underlying binary representation. Our contributions are two-fold. First, we demonstrate that the new distance represents a linear combination of the Lee and Hamming distance and derive upper bounds on the size of the codes under this metric based on linear programming techniques. Second, we propose a number of code constructions which imply lower bounds