Swapnil Bhatia

Functional optimization of gene clusters by combinatorial design and assembly

Large microbial gene clusters encode useful functions, including energy utilization and natural product biosynthesis, but genetic manipulation of such systems is slow, difficult and complicated by complex regulation. We exploit the modularity of a refactored Klebsiella oxytoca nitrogen fixation (nif) gene cluster (16 genes, 103 parts) to build genetic permutations that could not be achieved by starting from the wild-type cluster. Constraint-based combinatorial design and DNA assembly are used to build libraries of radically different cluster architectures by varying part choice, gene order, gene orientation and operon occupancy. We construct 84 variants of the nifUSVWZM operon, 145 variants of the nifHDKY operon, 155 variants of the nifHDKYENJ operon and 122 variants of the complete 16-gene pathway. The performance and behavior of these variants are characterized by nitrogenase assay and strand-specific RNA sequencing (RNA-seq), and the results are incorporated into subsequent design cycles. We have produced a fully synthetic cluster that recovers 57% of wild-type activity. Our approach allows the performance of genetic parts to be quantified simultaneously in hundreds of genetic contexts. This parallelized design-build-test-learn cycle, which can access previously unattainable regions of genetic space, should provide a useful, fast tool for genetic optimization and hypothesis testing.

Mobile video delivery with HTTP

Expansion in 3G cellular coverage and the emergence of more powerful mobile devices has increased demand for massively scalable mobile video delivery. The rapid adoption of the third screen as a primary screen for video has highlighted inefficiencies in the mobile delivery ecosystem and scalability issues in the mobile delivery infrastructure. This article provides an overview of the current mobile content delivery ecosystem and discusses the expanding role of HTTP-based mobile video delivery. A new class of HTTP-based mobile delivery protocols seeks to address existing quality and scalability issues by simplifying and standardizing mobile video delivery. This article shows how segment-based delivery has enabled HTTP-based live streaming and dynamic bitrate adaptation while increasing scalability through the use of existing CDN infrastructure.

An End-to-End Workflow for Engineering of Biological Networks from High-Level Specifications

We present a workflow for the design and production of biological networks from high-level program specifications. The workflow is based on a sequence of intermediate models that incrementally translate high-level specifications into DNA samples that implement them. We identify algorithms for translating between adjacent models and implement them as a set of software tools, organized into a four-stage toolchain: Specification, Compilation, Part Assignment, and Assembly. The specification stage begins with a Boolean logic computation specified in the Proto programming language. The compilation stage uses a library of network motifs and cellular platforms, also specified in Proto, to transform the program into an optimized Abstract Genetic Regulatory Network (AGRN) that implements the programmed behavior. The part assignment stage assigns DNA parts to the AGRN, drawing the parts from a database for the target cellular platform, to create a DNA sequence implementing the AGRN. Finally, the assembly stage computes an optimized assembly plan to create the DNA sequence from available part samples, yielding a protocol for producing a sample of engineered plasmids with robotics assistance. Our workflow is the first to automate the production of biological networks from a high-level program specification. Furthermore, the workflows modular design allows the same program to be realized on different cellular platforms simply by swapping workflow configurations. We validated our workflow by specifying a small-molecule sensor-reporter program and verifying the resulting plasmids in both HEK 293 mammalian cells and in E. coli bacterial cells.

Automated Selection of Synthetic Biology Parts for Genetic Regulatory Networks

Raising the level of abstraction for synthetic biology design requires solving several challenging problems, including mapping abstract designs to DNA sequences. In this paper we present the first formalism and algorithms to address this problem. The key steps of this transformation are feature matching, signal matching, and part matching. Feature matching ensures that the mapping satisfies the regulatory relationships in the abstract design. Signal matching ensures that the expression levels of functional units are compatible. Finally, part matching finds a DNA part sequence that can implement the design. Our software tool MatchMaker implements these three steps.

Analysis of the Gated IPACT Scheme for EPONs

Interleaved Polling with Adaptive Cycle Time (IPACT) is one of the earliest proposed polling schemes for dynamic bandwidth allocation in Ethernet Passive Optical Networks (EPONs) and has been extensively used as a benchmark by many subsequent allocation schemes. In this paper, we attempt to construct a mathematical model of the IPACT scheme under the gated service discipline. For N = 1 ONU, we derive closed-form expression for the steady state grant size. For N > 1 ONUs, we need to consider separately a small and a large load-distance ratio. For the former case, the N = 1 ONU model holds even for N > 1. For the latter case, we find a closed form expression for the grant size. Our model shows a reasonable match with the values obtained from simulation for the steady state queue size and hence the throughput and delay.

Pigeon: A Design Visualizer for Synthetic Biology

Pigeon is a Web-based tool that translates a textual description of a synthetic biology design into an image. It allows programmatic generation of design visualizations, is easy to learn, is easily extensible to new glyphs and notation, and can be connected to other software tools for visualizing their output. We present the Pigeon syntax, its current command set, and some examples of Pigeon programs and their output.

SBOL Visual: A Graphical Language for Genetic Designs.

Synthetic Biology Open Language (SBOL) Visual is a graphical standard for genetic engineering. It consists of symbols representing DNA subsequences, including regulatory elements and DNA assembly features. These symbols can be used to draw illustrations for communication and instruction, and as image assets for computer-aided design. SBOL Visual is a community standard, freely available for personal, academic, and commercial use (Creative Commons CC0 license). We provide prototypical symbol images that have been used in scientific publications and software tools. We encourage users to use and modify them freely, and to join the SBOL Visual community: http://www.sbolstandard.org/visual.

Constrained De Novo Sequencing of Conotoxins

De novo peptide sequencing by mass spectrometry (MS) can determine the amino acid sequence of an unknown peptide without reference to a protein database. MS-based de novo sequencing assumes special importance in focused studies of families of biologically active peptides and proteins, such as hormones, toxins, and antibodies, for which amino acid sequences may be difficult to obtain through genomic methods. These protein families often exhibit sequence homology or characteristic amino acid content; yet, current de novo sequencing approaches do not take advantage of this prior knowledge and, hence, search an unnecessarily large space of possible sequences. Here, we describe an algorithm for de novo sequencing that incorporates sequence constraints into the core graph algorithm and thereby reduces the search space by many orders of magnitude. We demonstrate our algorithm in a study of cysteine-rich toxins from two cone snail species (Conus textile and Conus stercusmuscarum) and report 13 de novo and about 60 total toxins.

IPACT with Smallest Available Report First: A New DBA Algorithm for EPON

Dynamic Bandwidth allocation in Ethernet Passive Optical Networks (EPONs) has been an area of intense research in recent years. Most of the proposed solutions offer clever methods for fair grant sizing, traffic prediction, and prioritized, differentiated services. Barring some work by Kamal et al. and some elements in the scheme proposed by Ma et al., no work has been done on exploring the order of granting (i.e., ONU sequencing) in an EPON. In this paper, we propose an unexplored heuristic for improving the performance of the IPACT scheme with respect to the most important metric: packet delay. In this heuristic, the OLT always grants that ONU which has the Smallest (Available) Reported queue length, First (SARF). Our simulations indicate that our heuristic can improve the delay performance of IPACT by 10-20% (when tested under the gated allocation policy).

Synthetic Biology Open Language Visual (SBOL Visual) Version 2.0

Abstract People who are engineering biological organisms often find it useful to communicate in diagrams, both about the structure of the nucleic acid sequences that they are engineering and about the functional relationships between sequence features and other molecular species. Some typical practices and conventions have begun to emerge for such diagrams. The Synthetic Biology Open Language Visual (SBOL Visual) has been developed as a standard for organizing and systematizing such conventions in order to produce a coherent language for expressing the structure and function of genetic designs. This document details version 2.0 of SBOL Visual, which builds on the prior SBOL Visual 1.0 standard by expanding diagram syntax to include functional interactions and molecular species, making the relationship between diagrams and the SBOL data model explicit, supporting families of symbol variants, clarifying a number of requirements and best practices, and significantly expanding the collection of diagram glyphs.