Virtual in situs: Sequencing mRNA from cryo-sliced Drosophila embryos to determine genome-wide spatial patterns of gene expression

Complex spatial and temporal patterns of gene expression underlie embryo differentiation, yet methods do not yet exist for the efficient genome-wide determination of spatial expression patterns during development. In situ imaging of transcripts and proteins is the gold-standard, but it is difficult and time consuming to apply to an entire genome, even when highly automated. Sequencing, in contrast, is fast and genome-wide, but is generally applied to homogenized tissues, thereby discarding spatial information. It is likely that these methods will ultimately converge, and we will be able to sequence RNAs in situ, simultaneously determining their identity and location. As a step along this path, we developed methods to cryosection individual blastoderm stage Drosophila melanogaster embryos along the anterior-posterior axis and sequence the mRNA isolated from each 25 micron slice. The spatial patterns of gene expression we infer closely match patterns previously determined by in situ hybridization and microscopy. We applied this method to generate a genome-wide timecourse of spatial gene expression from shortly after fertilization through gastrulation. We identify numerous genes with spatial patterns that have not yet been described in the several ongoing systematic in situ based projects. This simple experiment demonstrates the potential for combining careful anatomical dissection with high-throughput sequencing to obtain spatially resolved gene expression on a genome-wide scale.Comment: 6 pages, 3 figures, 7 supplemental figures (available on request from [email protected]

From Prosecutorial to Reparatory: A Valuable Post-Conflict Change of Focus

The ICC is well known in international legal circles. Indeed, everyone who knows anything about international law knows that the ICC is the acronym for the International Criminal Court, the body charged with prosecuting international crimes around the globe. Created in 2002, the ICC was intended to “put an end to impunity” for the perpetrators of international crimes” and to affirm “that the most serious crimes of concern to the international community as a whole must not go unpunished.”1 Imagine, however, a world where the “ICC” instead was an acronym for the International Compensation Court. That is, what if the ICC were a body charged with providing financial reparations to victims of mass violence? What if, instead of devoting millions upon millions of dollars to prosecuting those who commit international crimes, the international community used those resources to compensate victims of international crimes

Verification of Autonomous Systems: Developmental Test and Evaluation of an Autonomous UAS Swarming Algorithm Combining Simulation, Formulation and Live Flight

This research was driven by the increase of autonomous systems in the current millennium and the challenging nature of testing and evaluating their performance. A review of the current literature revealed proposed methods for verifying autonomous systems, but few implementations. It exposed several gaps in the current verification and validation methods and suggested goals for filling them. Through the use of modeling, software in the loop (SITL), and flight test, this research verified an autonomous swarming algorithm for unmanned aerial systems (UAS) and validated an exemplar of a testing framework. Thirteen sets of three-vehicle swarm data produced over two days of flight testing provided a baseline algorithm analysis. During these tests, vehicle separation distances deviated an average of 5.61 meters from the ideal state, with separation distance violations \u3c 6:39% of the time. The swarm achieved a 0.27 m average deviation and 0.43% violation in the best cases. Average packet loss between vehicles was 4.94% at a 5 Hz update rate, with an optimal communication lag \u3c 0:04 seconds. The multi-faceted empirical analysis created through the pairing of qualitative and quantitative analysis provided a complete understanding of vehicle behavior. This analysis also identified various areas of improvement for the algorithm and testing framework. The outcomes of this research formed a baseline testing continuum that is adaptable to a variety of follow-on investigations into formal verification of autonomous systems

Human Trafficking: Empowering Healthcare Providers and Community Partners as Advocates for Victims

Human trafficking (HT) is a public health crisis and the need for education is dire. Healthcare providers lack confidence in victim recognition and aid provision all while encountering victims in their practice. The project leader aimed to empower healthcare providers in recognizing signs of HT and making appropriate referrals. Ten community volunteers, including registered nurses, who assist with a mobile medical unit and spiritual 12-step program, participated in a Human Trafficking 101 training. The application of holistic nursing core principles and Human Caring Science was integrated as vital in victim identification. A pretest posttest design was utilized to administer the Perceived Competence Scale. A significant increase from pretest to posttest scores was found (t(9) = -6.567, p \u3c .05) in HT risk and signs identification, making referrals, and screening tool use. The participants also maintained a log documenting the number of times the screening tool and/or education gained from the training were used over a three-month period. The log did not reveal any identified victims at the conclusion of the three-month period. A well-designed training can significantly increase knowledge in medical and non-medical audiences. Nurses should extend training beyond traditional healthcare settings by reaching volunteers serving vulnerable populations

Procuring Guilty Pleas for International Crimes: The Limited Influence of Sentence Discounts

Approximately 90 percent of all American criminal cases are disposed of by means of guilty pleas, and a large percentage of defendants brought before courts in England, Australia, and other countries that use common-law procedures likewise plead guilty. Why do substantial numbers of defendants in national criminal justice systems choose to convict themselves when they are entitled to have their guilt formally adjudicated? The widely accepted primary reason is that they receive sentencing discounts when they choose to selfconvict. Most defendants charged with domestic crimes plead guilty following a process of plea bargaining between defense counsel and prosecutors. Although plea bargaining can take many forms, at its heart is a promise of some form of sentence leniency in exchange for the defendant\u27s guilty plea. In the context of domestic crimes, then, most defendants are understood to plead guilty primarily, if not exclusively, in order to obtain sentence discounts, and the magnitude of the available discounts will largely determine how many guilty pleas will be obtained

Genetic Tools and Ecological Context for Commensal Rodent Management: Population Genetic Analysis and Gene Drives

Invasive commensal rodents are the source of significant harms across diverse ecosystems globally, including biodi­versity loss on islands, economic damage in agricultural settings, and disease spread in urban areas. Genetic tools can provide unique insights and solutions for rodent management or eradication, adding to the toolbox of integrated pest management opera­tions. In real world conditions, the utility of genetic tools for rodent management is constrained by ecological factors such as mating and dispersal behavior, landscape variation, and ongoing management strategies. Here we describe two distinct uses of genetic tools for rodent management and their ecological considerations. First, we describe how patterns of genetic connectivity can inform the establishment of functional management units, discern among scenarios leading to island eradication failure, and provide insights into rodent management outcomes. Second, we discuss the utility and risks of using gene drive systems to eradicate invasive house mice from island environments. We describe aspects of risk assessment and mitigation as well as the utility of computational models for improving gene drive preparedness. Together, we outline the importance of ecologically-informed implementation strategies when using genetic tools, both currently available and in development, for the management of commensal rodents

Testing validation tools on CLIPS-based expert systems

The Expert Systems Validation Associate (EVA) is a validation system which was developed at the Lockheed Software Technology Center and Artificial Intelligence Center between 1986 and 1990. EVA is an integrated set of generic tools to validate any knowledge-based system written in any expert system shell such as C Language Integrated Production System (CLIPS), ART, OPS5, KEE, and others. Many validation tools have been built in the EVA system. In this paper, we describe the testing results of applying the EVA validation tools to the Manned Maneuvering Unit (MMU) Fault Diagnosis, Isolation, and Reconfiguration (FDIR) expert system, written in CLIPS, obtained from the NASA Johnson Space Center