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101 Total Publications
HOME | RESOURCES | Publications


101 Total Publications


TITLES and AUTHORS Year Link PDF + Abstract
  • Synthesis, debugging, and effects of synthetic chromosome consolidation: synVI and beyond Leslie A. Mitchell, et. al.

    Institution: NYU School of Medicine, Boeke Lab

    Publication: Science

    2017 PDF abstract

    Total synthesis of designer chromosomes and genomes is a new paradigm for the study of genetics and biological systems. The Sc2.0 project is building a designer yeast genome from scratch to test and extend the lim- its of our biological knowledge. Here we describe the design, rapid assembly, and characterization of synthetic chromosome VI (synVI). Further, we investigate the phenotypic, transcriptomic, and some consolidation to uncover possible synthetic genetic interactions and/or perturbations of native cellular networks as the number of designer changes increases is the next major step for the Sc2.0 project.

    Publication / Type:
    Related Subject:
    Leslie A. Mitchell, Synthetic Biology, Genomics
  • Bug mapping and fitness testing of chemically synthesized chromosome X Yi Wu, et. al.

    Institution: NYU School of Medicine, Boeke Lab

    Publication: Science

    2017 PDF abstract

    Design and construction of an extensively modified yeast genome is a direct means to interrogate the integrity, com- prehensiveness, and accuracy of the knowl- edge amassed by the yeast community to date. The international synthetic yeast genome project (Sc2.0) aims to build an entirely de- signer, synthetic Saccharomyces cerevisiae ge- nome. The synthetic genome is designed to increase genome stability and genetic flexi- bility while maintaining cell fitness near that of the wild type. A major challenge for a ge- nome synthesis lies in identifying and elim- inating fitness-reducing sequence variants referred to as “bugs.”

    Publication / Type:
    Related Subject:
    Yi Wu, Synthetic Biology, Genomics
  • Precision Cancer Medicine in the Acoustic Dispensing Era: Ex Vivo Primary Cell Drug Sensitivity Testing Evgeny Kulesskiy, Jani Saarela, Laura Turunen, and Krister Wennerberg

    Institution: Institute for Molecular Medicine Finland (FIMM), University of Helsinki, Finland

    Publication: Journal of Laboratory Automation (JALA) Special Issue

    2016 PDF abstract

    Cancer therapy is increasingly becoming individualized, but there are also big gaps between the molecular knowledge of individual cancers we can generate today and what can be applied in the clinic. In an attempt to bridge this knowledge gap between cancer genetic and molecular profiling and clinically useful information, an individualized systems medicine program has been established at the Institute for Molecular Medicine Finland (FIMM), University of Helsinki, and the Helsinki University Hospital. Central to this program is drug sensitivity and resistance testing (DSRT), in which responses of primary cancer cells to a comprehensive clinical oncology and signal transduction drug collection are monitored. The drug sensitivity information is used with molecular profiling to establish hypotheses on individual cancer-selective targeting drug combinations and their predictive biomarkers, which can be explored in the clinic. Here, we describe how acoustic droplet ejection is enabling DSRT in our cancer individualized systems medicine program to (1) generate consistent but configurable assay-ready plates and determine how this affects data quality, (2) flexibly prepare drug combination testing plates, (3) dispense reagents and cells to the assay plates, and (4) perform ultra-miniaturized follow up assays on the cells from DSRT plates.

    Publication / Type:
    Journal of Laboratory Automation (JALA) Special Issue
    Related Subject:
    automated biology, statistics, informatics and software, compound management, acoustic droplet ejection
  • Identification of b-hematin inhibitors in the MMV Malaria Box Fong K., Sandlin R. and Wright D.

    Institution: Vanderbilt University

    Publication: International Journal for Parasitology: Drugs and Drug Resistance, Volume 5, Issue 3, December 2015

    2015 abstract

    The Malaria Box, assembled by the Medicines for Malaria Venture, is a set of 400 structurally diverse, commercially available compounds with demonstrated activity against blood-stage Plasmodium falciparum. The compounds are a representative subset of the 20,000 in vitro antimalarials identified from the high-throughput screening efforts of St. Jude Children's Research Hospital (TN, USA), Novartis and GlaxoSmithKline. In addition, a small set of active compounds from commercially available libraries was added to this group, but it has not previously been published. Elucidation of the biochemical pathways on which these compounds act is a major challenge; therefore, access to these compounds has been made available free of charge to the investigator community. Here, the Malaria Box compounds were tested for activity against the formation of b-hematin, a synthetic form of the heme detoxification biomineral, hemozoin. Further, the mechanism of action of these compounds within the malaria parasite was explored. Ten of the Malaria Box compounds demonstrated significant inhibition of b-hematin formation. In this assay, doseeresponse data revealed IC50 values ranging from 8.7 to 22.7 mM for these hits, each of which is more potent than chloroquine (a known inhibitor of hemozoin formation). The in vitro antimalarial activity of these ten hits was confirmed in cultures of the chloroquine sensitive D6 strain of the parasite resulting in IC50 values of 135e2165 nM, followed by testing in the multidrug resistant strain, C235. Cultures of P. falciparum (D6) were then examined for their heme distribution following treatment with nine of the commercially available confirmed compounds, seven of which disrupted the hemozoin pathway.

    Publication / Type:
    International Journal for Parasitology: Drugs and Drug Resistance, Volume 5, Issue 3, December 2015
    Related Subject:
    b-hematin inhibitors, MMV malaria
  • Compound Transfer by Acoustic Droplet Ejection Promotes Quality and Efficiency in Ultra-High-Throughput Screening Campaigns Dawes T., Turincio R., Jones S., Rodriguez R., Gadiagellan D., Thana P., Clark K., Gustafson A., Orr

    Institution: Genentech

    Publication: Journal of Laboratory Automation (JALA) Special Issue

    2015 abstract

    Acoustic droplet ejection (ADE) as a means of transferring library compounds has had a dramatic impact on the way in which high-throughput screening campaigns are conducted in many laboratories. Two Labcyte Echo ADE liquid handlers form the core of the compound transfer operation in our 1536-well based ultra-high-throughput screening (uHTS) system. Use of these instruments has promoted flexibility in compound formatting in addition to minimizing waste and eliminating compound carryover. We describe the use of ADE for the generation of assay-ready plates for primary screening as well as for follow-up dose-response evaluations. Custom software has enabled us to harness the information generated by the ADE instrumentation. Compound transfer via ADE also contributes to the screening process outside of the uHTS system. A second fully automated ADE-based system has been used to augment the capacity of the uHTS system as well as to permit efficient use of previously picked compound aliquots for secondary assay evaluations. Essential to the utility of ADE in the high-throughput screening process is the high quality of the resulting data. Examples of data generated at various stages of high-throughput screening campaigns are provided. Advantages and disadvantages of the use of ADE in high-throughput screening are discussed.

    Publication / Type:
    Journal of Laboratory Automation (JALA) Special Issue
    Related Subject:
    high-throughput screening, acoustic droplet ejection, assay-ready plates
  • Delivering an Automated and Integrated Approach to Combination Screening Using Acoustic-Droplet Technology Cross K, Craggs R, Swift D, Stiaram A, Daya S, Roberts M, Hawley S, Owen P, and Isherwood V

    Institution: AstraZeneca and Tessela plc.

    Publication: J Lab Autom. 2015 Apr 2. DOI: 10.1177/2211068215579163

    2015 PDF abstract

    Drug combination testing in the pharmaceutical industry has typically been driven by late-stage opportunistic strategies rather than by early testing to identify drug combinations for clinical investigation that may deliver improved efficacy. A rationale for combinations exists across a number of diseases in which pathway redundancy or resistance to therapeutics are evident. However, early assays are complicated by the absence of both assay formats representative of disease biology and robust infrastructure to screen drug combinations in a medium-throughput capacity. When applying drug combination testing studies, it may be difficult to translate a study design into the required well contents for assay plates because of the number of compounds and concentrations involved. Dispensing these plates increases in difficulty as the number of compounds and concentration points increase and compounds are subsequently rolled onto additional labware. We describe the development of a software tool, in conjunction with the use of acoustic droplet technology, as part of a compound management platform, which allows the design of an assay incorporating combinations of compounds. These enhancements to infrastructure facilitate the design and ordering of assay-ready compound combination plates and the processing of combinations data from high-content organotypic assays.

    Publication / Type:
    J Lab Autom. 2015 Apr 2. DOI: 10.1177/2211068215579163
    Related Subject:
    laboratory informatics, combinations, acoustic droplet technology, apothecary, assay-ready plate, high-content screening, sample management
  • qPCRTag Analysis - A High Throughput, Real Time PCR Assay for Sc2.0 Genotyping Mitchell L, Phillips N, Lafont A, Martin J, Cutting R, and Boeke J

    Institution: Department of Biochemistry and Molecular Pharmacology, Institute for Systems Genetics, New York Univ

    Publication: J. Vis. Exp. (99), e52941, doi:10.3791/52941 (2015)

    2015 abstract

    The Synthetic Yeast Genome Project (Sc2.0) aims to build 16 designer yeast chromosomes and combine them into a single yeast cell. To date one synthetic chromosome, synIII1, and one synthetic chromosome arm, synIXR2, have been constructed and their in vivo function validated in the absence of the corresponding wild type chromosomes. An important design feature of Sc2.0 chromosomes is the introduction of PCRTags, which are short, re-coded sequences within open reading frames (ORFs) that enable differentiation of synthetic chromosomes from their wild type counterparts. PCRTag primers anneal selectively to either synthetic or wild type chromosomes and the presence/absence of each type of DNA can be tested using a simple PCR assay. The standard readout of the PCRTag assay is to assess presence/absence of amplicons by agarose gel electrophoresis However, with an average PCRTag amplicon density of one per 1.5 kb and a genome size of ~12 Mb, the completed Sc2.0 genome will encode roughly 8,000 PCRTags. To improve throughput, we have developed a real time PCR-based detection assay for PCRTag genotyping that we call qPCRTag analysis. The workflow specifies 500 nl reactions in a 1,536 multiwell plate, allowing us to test up to 768 PCRTags with both synthetic and wild type primer pairs in a single experiment.

    Publication / Type:
    J. Vis. Exp. (99), e52941, doi:10.3791/52941 (2015)
    Related Subject:
    Echo® Liquid Handler for OMICS , Echo® Liquid Handler for Screening and OMICS
  • Low-Cost, High-Throughput Sequencing of DNA Assemblies Using a Highly Multiplexed Nextera Process E Shapland, et. al.

    Institution: Amyris, Inc. and TOTAL New Energies USA, Inc.

    Publication: ACS Synth. Biol., Article ASAP DOI: 10.1021/sb500362n

    2015 abstract

    In recent years, next-generation sequencing (NGS) technology has greatly reduced the cost of sequencing whole genomes, whereas the cost of sequence verification of plasmids via Sanger sequencing has remained high. Consequently, industrial-scale strain engineers either limit the number of designs or take short cuts in quality control. Here, we show that over 4000 plasmids can be completely sequenced in one Illumina MiSeq run for less than $3 each (15× coverage), which is a 20-fold reduction over using Sanger sequencing (2× coverage). We reduced the volume of the Nextera tagmentation reaction by 100-fold and developed an automated workflow to prepare thousands of samples for sequencing. We also developed software to track the samples and associated sequence data and to rapidly identify correctly assembled constructs having the fewest defects. As DNA synthesis and assembly become a centralized commodity, this NGS quality control (QC) process will be essential to groups operating high-throughput pipelines for DNA construction.

    Publication / Type:
    ACS Synth. Biol., Article ASAP DOI: 10.1021/sb500362n
    Related Subject:
    synthetic biology; next-generation sequencing; NGS; high throughput, DNA Assembly, Sequencing, Echo® Liquid Handler for Screening and OMICS
  • Axitinib Effectively Inhibits BCR-ABL1(T315I) with a Distinct Binding Confirmation Pemovska T., Johnson E., Kontro M., Repasky G., Chen J., Wells P, Ciaran N., Cronin C., McTigue M.,

    Institution: Institute for Molecular Medicine Finland; University of Helsinki; La Jolla Laboratories, Pfizer

    Publication: Nature 519, 102-105 (March 5, 2015), doi:10.1038/nature14119

    2015 abstract

    The BCR-ABL1 fusion gene is a driver oncogene in chronic myeloid leukaemia and 30–50% of cases of adult acute lymphoblastic leukaemia1. Introduction of ABL1 kinase inhibitors (for example, imatinib) has markedly improved patient survival2, but acquired drug resistance remains a challenge3, 4, 5. Point mutations in the ABL1 kinase domain weaken inhibitor binding6 and represent the most common clinical resistance mechanism...

    Publication / Type:
    Nature 519, 102-105 (March 5, 2015), doi:10.1038/nature14119
    Related Subject:
    Translational research, oncogene
  • A Personalised Medicine Drug Sensitivity and Resistance Testing Platform and Utilisation of Acoustic Droplet Ejection at the Institute for Molecular Medicine Finland Saarela J, Kulesskiy E, Laamanen K, Pemovska T, Pietiainen V, Potdar S, von Schantz-Fant C, Turunen

    Institution: Institute for Molecular Medicine Finland, University of Helsinki, Finland (FIMM)

    Publication: Synergy Vol 1, Issue 1, Sep 2014, Pages 78 doi:10.1016/j.synres.2014.07.001

    2015 abstract

    The High Throughput Biomedicine (HTB) unit at FIMM Technology Centre provides a wide range of biomedical high throughput assays. In collaboration with research groups and the Hospital District of Helsinki and Uusimaa, we have set up drug sensitivity and resistance testing (DSRT) platform with a set of 450 approved and investigational oncology drugs on patient samples. Ex vivo drug testing is currently performed at FIMM with primary cancer cells from patients with leukaemia and multiple myeloma as well as cancer cell lines and the drug sensitivity responses are integrated with molecular profiling such as exome sequencing, transcriptomics and phosphoproteomics. Currently, DSRT is run in 384-well plate format in 5 concentrations for each drug or in 1536-well plate format with 9 concentrations per drug and the assay plates are pre-drugged using acoustic droplet ejection with Labcyte® Echo® 550. The viability and cell death of cells is measured after 72 h and results are analysed using Dotmatics Studies – software and an in-house developed interface, Breeze. Application of the platform to AML patient samples has uncovered taxonomic drug-response subtypes and individualised therapy based on DSRT has resulted in several clinical responses. The DSRT platform enables drug repositioning, provides new combinatorial possibilities and allows for linking drug sensitivities to predictive biomarkers. We are developing the platform with additional readouts and increasing the number of drugs. Our Labcyte Access Workstation, with Echo 550 and Echo® 525 integration, allows development and set-up of miniaturised follow-up assays, such as reverse-phase protein array and qPCR, using non-contact acoustic dispensing. 

    Publication / Type:
    Synergy Vol 1, Issue 1, Sep 2014, Pages 78 doi:10.1016/j.synres.2014.07.001
    Related Subject:
    Medicine Drug Sensitivity Resistance Testing Platform Utilisation Acoustic Droplet Ejection Institute of Molecular Medicine Finland

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