A Rapid FACS-Based Strategy to Isolate Human Gene Knockin and Knockout Clones

Gene targeting protocols for mammalian cells remain inefficient and labor intensive. Here we describe FASTarget, a rapid, fluorescent cell sorting based strategy to isolate rare gene targeting events in human somatic cells. A fluorescent protein is used as a means for direct selection of targeted clones obviating the need for selection and outgrowth of drug resistant clones. Importantly, the use of a promoter-less, ATG-less construct greatly facilitates the recovery of correctly targeted cells. Using this method we report successful gene targeting in up to 94% of recovered human somatic cell clones. We create functional EYFP-tagged knockin clones in both transformed and non-transformed human somatic cell lines providing a valuable tool for mammalian cell biology. We further demonstrate the use of this technology to create gene knockouts. Using this generally applicable strategy we can recover gene targeted clones within approximately one month from DNA construct delivery to obtaining targeted monoclonal cell lines.

Second St. Gallen European Organisation for Research and Treatment of Cancer Gastrointestinal Cancer Conference: consensus recommendations on controversial issues in the primary treatment of rectal cancer

Primary treatment of rectal cancer was the focus of the second St. Gallen European Organisation for Research and Treatment of Cancer (EORTC) Gastrointestinal Cancer Conference. In the context of the conference, a multidisciplinary international expert panel discussed and voted on controversial issues which could not be easily answered using published evidence. Main topics included optimal pretherapeutic imaging, indication and type of neoadjuvant treatment, and the treatment strategies in advanced tumours. Here we report the key recommendations and summarise the related evidence. The treatment strategy for localised rectal cancer varies from local excision in early tumours to neoadjuvant radiochemotherapy (RCT) in combination with extended surgery in locally advanced disease. Optimal pretherapeutic staging is a key to any treatment decision. The panel recommended magnetic resonance imaging (MRI) or MRI + endoscopic ultrasonography (EUS) as mandatory staging modalities, except for early T1 cancers with an option for local excision, where EUS in addition to MRI was considered to be most important because of its superior near-field resolution. Primary surgery with total mesorectal excision was recommended by most panellists for some early tumours with limited risk of recurrence (i.e. cT1-2 or cT3a N0 with clear mesorectal fascia on MRI and clearly above the levator muscles), whereas all other stages were considered for multimodal treatment. The consensus panel recommended long-course RCT over short-course radiotherapy for most clinical situations where neoadjuvant treatment is indicated, with the exception of T3a/b N0 tumours where short-course radiotherapy or even no neoadjuvant therapy were regarded to be an option. In patients with potentially resectable tumours and synchronous liver metastases, most panel members did not see an indication to start with classical fluoropyrimidine-based RCT but rather favoured preoperative short-course radiotherapy with systemic combination chemotherapy or alternatively a liver-first resection approach in resectable metastases, which both allow optimal systemic therapy for the metastatic disease. In general, proper patient selection and discussion in an experienced multidisciplinary team was considered as crucial component of care.

Differences in male coloration are predicted by divergent sexual selection between populations of a cichlid fish

Female mating preferences can influence both intraspecific sexual selection and interspecific reproductive isolation, and have therefore been proposed to play a central role in speciation. Here, we investigate experimentally in the African cichlid fish Pundamilia nyererei if differences in male coloration between three para-allopatric populations (i.e. island populations with gene flow) of P. nyererei are predicted by differences in sexual selection by female mate choice between populations. Second, we investigate if female mating preferences are based on the same components of male coloration and go in the same direction when females choose among males of their own population, their own and other conspecific populations and a closely related para-allopatric sister-species, P. igneopinnis. Mate-choice experiments revealed that females of the three populations mated species-assortatively, that populations varied in their extent of population-assortative mating and that females chose among males of their own population based on different male colours. Females of different populations exerted directional intrapopulation sexual selection on different male colours, and these differences corresponded in two of the populations to the observed differences in male coloration between the populations. Our results suggest that differences in male coloration between populations of P. nyererei can be explained by divergent sexual selection and that population-assortative mating may directly result from intrapopulation sexual selection.

Sea ice habitat characteristics and number of narwhal (Monodon monoceros) sightings in Disko Bay, West Greenland

There is a paucity of information on abundance, densities, and habitat selection of narwhals Monodon monoceros in the offshore pack ice of Baffin Bay, West Greenland, despite the critical importance of winter foraging regions and considerable sea ice declines in the past decades. We conducted a double-platform visual aerial survey over a narwhal wintering ground to obtain pack ice densities and develop the first fully corrected abundance estimate using point conditional mark-recapture distance sampling. Continuous video recording and digital images taken along the trackline allowed for in situ quantification of winter narwhal habitat and for the estimation of fine-scale narwhal habitat selection and habitat-specific sighting probabilities. Abundance at the surface was estimated at 3484 (coefficient of variation [CV] = 0.46) including whales missed by observers. The fully corrected abundance of narwhals was 18 044 (CV = 0.46), or approximately one-quarter of the entire Baffin Bay population. The narwhal wintering ground surveyed (~9500 km**2) had 2.4 to 3.2% open water based on estimates from satellite imagery (NASA Moderate Resolution Imaging Spectroradiometer) and 1565 digital photographic images collected on the trackline. Thus, the ~18 000 narwhals had access to 233 km**2 of open water, resulting in an average density of ~77 narwhals/km**2 open water. Narwhal sighting probability near habitats with <10% or 10 to 50% open water was significantly higher than sighting probability in habitats with >50% open water, suggesting narwhals select optimal foraging areas in dense pack ice regardless of open water availability. This study provides the first quantitative ecological data on densities and habitat selection of narwhals in pack ice foraging regions that are rapidly being altered with climate change.

Pharmaceutical industry : hazardous waste generation, treatment, and disposal.

Final report (SW-508) describing work performed for the Federal Solid waste management program under Contract no. 68-01-2684.

The use of computers in literature searching and related reference activities in libraries : papers presented at the 1975 Clinic on Library Applications of Data Processing, April 27-30, 1975 /

Includes bibliographical references.

Solid waste handling and disposal in multistory buildings and hospitals.

"Research initiated by the Project Planning and Pollution Control Division, Dept. of County Engineer, County of Los Angeles."

Solid waste handling and disposal in multistory buildings and hospitals.

"Research initiated by the Project Planning and Pollution Control Division, Dept. of County Engineer, County of Los Angeles."

Combined desalination and power generation using coal energy.

Phase I report. Site selection and estimated water costs based on current technology. Phase II report. Investigation of emerging technology, by-product recovery, and very large capacities. Period of performance: June 1965-Oct. 1966.

Replication of Zeidner, Johnson, and colleagues' method for estimating Army aptitude area (AA) composites /

"Selection and Assignment Research Unit."

Essays and addresses,

Published in commemoration of the opening of the new college buildings, October 7th, 1873.

Essays on Personal Bankruptcy Law and Small Business Financing

Thesis (Ph.D.)--University of Washington, 2016-06

Submodular Optimization and Data Processing

Thesis (Ph.D.)--University of Washington, 2016-06

Remotely sensed data for ecosystem analyses: Combining hierarchy and scene models

Remotely sensed data have been used extensively for environmental monitoring and modeling at a number of spatial scales; however, a limited range of satellite imaging systems often. constrained the scales of these analyses. A wider variety of data sets is now available, allowing image data to be selected to match the scale of environmental structure(s) or process(es) being examined. A framework is presented for use by environmental scientists and managers, enabling their spatial data collection needs to be linked to a suitable form of remotely sensed data. A six-step approach is used, combining image spatial analysis and scaling tools, within the context of hierarchy theory. The main steps involved are: (1) identification of information requirements for the monitoring or management problem; (2) development of ideal image dimensions (scene model), (3) exploratory analysis of existing remotely sensed data using scaling techniques, (4) selection and evaluation of suitable remotely sensed data based on the scene model, (5) selection of suitable spatial analytic techniques to meet information requirements, and (6) cost-benefit analysis. Results from a case study show that the framework provided an objective mechanism to identify relevant aspects of the monitoring problem and environmental characteristics for selecting remotely sensed data and analysis techniques.

Orientation of the genetic variance-covariance matrix and the fitness surface for multiple male sexually selected traits

Stabilizing selection has been predicted to change genetic variances and covariances so that the orientation of the genetic variance-covariance matrix (G) becomes aligned with the orientation of the fitness surface, but it is less clear how directional selection may change G. Here we develop statistical approaches to the comparison of G with vectors of linear and nonlinear selection. We apply these approaches to a set of male sexually selected cuticular hydrocarbons (CHCs) of Drosophila serrata. Even though male CHCs displayed substantial additive genetic variance, more than 99% of the genetic variance was orientated 74.9degrees away from the vector of linear sexual selection, suggesting that open-ended female preferences may greatly reduce genetic variation in male display traits. Although the orientation of G and the fitness surface were found to differ significantly, the similarity present in eigenstructure was a consequence of traits under weak linear selection and strong nonlinear ( convex) selection. Associating the eigenstructure of G with vectors of linear and nonlinear selection may provide a way of determining what long-term changes in G may be generated by the processes of natural and sexual selection.