Brian C. Ross

Decoding long nanopore sequencing reads of natural DNA

Nanopore sequencing of DNA is a single-molecule technique that may achieve long reads, low cost and high speed with minimal sample preparation and instrumentation. Here, we build on recent progress with respect to nanopore resolution and DNA control to interpret the procession of ion current levels observed during the translocation of DNA through the pore MspA. As approximately four nucleotides affect the ion current of each level, we measured the ion current corresponding to all 256 four-nucleotide combinations (quadromers). This quadromer map is highly predictive of ion current levels of previously unmeasured sequences derived from the bacteriophage phi X 174 genome. Furthermore, we show nanopore sequencing reads of phi X 174 up to 4,500 bases in length, which can be unambiguously aligned to the phi X 174 reference genome, and demonstrate proof-of-concept utility with respect to hybrid genome assembly and polymorphism detection. This work provides a foundation for nanopore sequencing of long, natural DNA strands.

Subangstrom single-molecule measurements of motor proteins using a nanopore

Techniques for measuring the motion of single motor proteins, such as FRET and optical tweezers, are limited to a resolution of ∼300 pm. We use ion current modulation through the protein nanopore MspA to observe translocation of helicase Hel308 on DNA with up to ∼40 pm sensitivity. This approach should be applicable to any protein that translocates on DNA or RNA, including helicases, polymerases, recombinases and DNA repair enzymes.

Mutual Information between Discrete and Continuous Data Sets

Mutual information (MI) is a powerful method for detecting relationships between data sets. There are accurate methods for estimating MI that avoid problems with “binning” when both data sets are discrete or when both data sets are continuous. We present an accurate, non-binning MI estimator for the case of one discrete data set and one continuous data set. This case applies when measuring, for example, the relationship between base sequence and gene expression level, or the effect of a cancer drug on patient survival time. We also show how our method can be adapted to calculate the Jensen–Shannon divergence of two or more data sets.

Systematic Analysis of Pleiotropy in C. elegans Early Embryogenesis

Pleiotropy refers to the phenomenon in which a single gene controls several distinct, and seemingly unrelated, phenotypic effects. We use C. elegans early embryogenesis as a model to conduct systematic studies of pleiotropy. We analyze high-throughput RNA interference (RNAi) data from C. elegans and identify “phenotypic signatures”, which are sets of cellular defects indicative of certain biological functions. By matching phenotypic profiles to our identified signatures, we assign genes with complex phenotypic profiles to multiple functional classes. Overall, we observe that pleiotropy occurs extensively among genes involved in early embryogenesis, and a small proportion of these genes are highly pleiotropic. We hypothesize that genes involved in early embryogenesis are organized into partially overlapping functional modules, and that pleiotropic genes represent “connectors” between these modules. In support of this hypothesis, we find that highly pleiotropic genes tend to reside in central positions in protein-protein interaction networks, suggesting that pleiotropic genes act as connecting points between different protein complexes or pathways.

Halting the Revolving Door of Faculty Turnover: Recruiting and Retaining Clinician Educators in an Academic Medical Simulation Center

Simulation-based education is indispensable in preparing healthcare providers for patient care.1 Simulation centers and programs that serve as a critical platform for promoting patient safety and high-quality training depend on multiple requirements for success: diversified and sustainable financing,2–5 technical personnel with a long-term commitment to simulation education,6,7 simulation and information technology infrastructure designed to match priority training needs,8–12 and resources for curricular development, instruction, faculty development, and research.6,7,9,12–14 An additional requirement not widely discussed in the literature is the recruitment and retention of faculty who serve as simulation educators, which is the focus of this report.

Direct Detection of Unnatural DNA Nucleotides dNaM and d5SICS using the MspA Nanopore

Malyshev et al. showed that the four-letter genetic code within a living organism could be expanded to include the unnatural DNA bases dNaM and d5SICS. However, verification and detection of these unnatural bases in DNA requires new sequencing techniques. Here we provide proof of concept detection of dNaM and d5SICS in DNA oligomers via nanopore sequencing using the nanopore MspA. We find that both phi29 DNA polymerase and Hel308 helicase are capable of controlling the motion of DNA containing dNaM and d5SICS through the pore and that single reads are sufficient to detect the presence and location of dNaM and d5SICS within single molecules.

Randomised trial comparing the recording ability of a novel, electronic emergency documentation system with the AHA paper cardiac arrest record.

Objective To evaluate the ability of an electronic system created at the University of Washington to accurately document prerecorded VF and pulseless electrical activity (PEA) cardiac arrest scenarios compared with the American Heart Association paper cardiac arrest record. Methods 16 anaesthesiology residents were randomly assigned to view one of two prerecorded, simulated VF and PEA scenarios and asked to document the event with either the paper or electronic system. Each subject then repeated the process with the other video and documentation method. Five types of documentation errors were defined: (1) omission, (2) specification, (3) timing, (4) commission and (5) noise. The mean difference in errors between the paper and electronic methods was analysed using a single factor repeated measures ANOVA model. Results Compared with paper records, the electronic system omitted 6.3 fewer events (95% CI −10.1 to −2.5, p=0.003), which represents a 28% reduction in omission errors. Users recorded 2.9 fewer noise items (95% CI −5.3 to −0.6, p=0.003) when compared with paper, representing a 36% decrease in redundant or irrelevant information. The rate of timing (Δ=−3.2, 95% CI −9.3 to 3.0, p=0.286) and commission (Δ=−4.4, 95% CI −9.4 to 0.5, p=0.075) errors were similar between the electronic system and paper, while the rate of specification errors were about a third lower for the electronic system when compared with the paper record (Δ=−3.2, 95% CI −6.3 to −0.2, p=0.037). Conclusions Compared with paper documentation, documentation with the electronic system captured 24% more critical information during a simulated medical emergency without loss in data quality.

Simopoly: improving simulation center operations through a tabletop simulation exercise.

Summary Statement Health care simulation is a rapidly growing, heterogeneous field requiring expertise that is not traditionally represented within health care or educational institutions. Simulation staff members often take on unique roles and have goals and career development needs that differ from more typical hospital and educational institution-based personnel. This poses both challenges and opportunities for simulation administrators. In this article, we describe a novel simulation, Simopoly, designed to provide the opportunity to think creatively about simulation center staff development, retention, and promotion in the context of daily operations and management.