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Increasing gene editing efficiencies in eukaryotic cell lines by selection of appropriate CRISPR-Cas9 reagents
Melissa L. Kelley, Žaklina Strezoska, Elena Maksimova, Hidevaldo Machado, Emily M. Anderson, Maren Mayer, Annaleen Vermeulen, Shawn McClelland, Anja van Brabant Smith

Overview of various CRISPR-Cas9 reagents to provide the highest efficiency of gene editing in your experiments.

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Knockdown of p53 by Accell self-delivering siRNA causes inhibition of p53-dependent DNA damage response in IMR-32 neuroblastoma cell line and β-amyloid toxicity in rat cortical neurons
Žaklina Strezoska, Tamara Seredenina1, Devin Leake, Annaleen Vermeulen

Here we describe how application of Accell siRNA enabled the development of a high content screening assay in IMR-32 neuroblastoma cells and a whole culture cell viability assay in primary rat cortical neurons.

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An Efficient Method for the Incorporation of Molecular Probes at Multiple/Specific sites in RNA: Levulinyl Protection for 2'-ACE ® , 5'-Silyl Oligoribonucleotide Synthesis
Xiaoqin Cheng, Shawn Begay, Randy Rauen, Kelly Grimsley, Kaizhang He, Michael Delaney

A unique method that uses a levulinate ester as a protecting group to introduce conjugates or molecular probes to virtually any location in a synthetic RNA molecule is discussed. The Levulinyl protecting group is stable in RNA synthesis conditions and can be removed without affecting the other parts of the synthesized RNA. We show the capabilities of this approach with three high-complexity synthesis examples.

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Innovative technology that enables RNAi in difficult to transfect cells
Christina Yamada, Kathryn Robinson, Allison St. Amand, Zaklina Strezoska, Greg Wardle, Anastasia Khvorova, Devin Leake

Investigations at Dharmacon have led to the development of innovative siRNA molecules that can be delivered into difficult-to-transfect cells without additional lipid reagents, virus, or instruments. This technology, Accell siRNA reagents, enables gene knockdown for functional genomic studies in a wide variety of cell types. In some instances, cells can be continuously dosed with Accell siRNAs to enable target gene knockdown for extended durations.

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Innovative technology that enables RNAi in difficult to transfect cells
Christina Yamada, Kathryn Robinson, Allison St. Amand, Zaklina Strezoska, Greg Wardle, Anastasia Khvorova, Devin Leake

Investigations at Dharmacon have led to the development of innovative siRNA molecules that can be delivered into difficult-to-transfect cells without additional lipid reagents, virus, or instruments. This technology, Accell siRNA reagents, enables gene knockdown for functional genomic studies in a wide variety of cell types. In some instances, cells can be continuously dosed with Accell siRNAs to enable target gene knockdown for extended durations.

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Modelling CLL cell and T-cell Migration in a Dynamic Circulating Model of CLL
Elisabeth Walsbyl, Paul Brennan', Guy Pratte, Andrea Buggins3, Tanja N Hartmann'', Chris Fegan1 and Chris Pepper'

We have recently developed a novel circulating model of chronic lymphocytic leukaemia (CLL) that mimics the transient interactions that take place between circulating lymphocytes and vascular endothelium. Here we show that both normal and malignant lymphocytes actively underwent transendothelial migration.

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Measuring Antitumor Effect of c-Myr-Max heterodimerization inhibitor 100258-F4 on Ovarian Cancer Cells using Cellometer Imaging Cytometry
Leo L. Chan, Jiandong Wang, Xiaoli Ma, Hannah M. Jones, Fang Song, Weiyuan Zhang, Victoria L. Bae-Jump, Chunxiao Zhou

The effect from the c-Mycinhibitor 100258-F4 was clearly observed from the G0/G1 cell cycle arrest and the increase in early and late apoptotic cell. The Cellometer method can measure fluorescent cell-based assays such as cell cycle, apoptosis, protein expression, autophagy and viability. The ability to export the data to FCS Express facilitates simple data analysis and reporting to generate results for publications.

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Quantification of Natural Killer Cell-Mediated Cytotixicity using Celigo Imaging Cytometry
Leo L. Chan, Srinivas S. Somanchi, Kelsey Rosbach, Dean A. Lee

Time-course tracking of % lysis eliminates multiple controls & the effect of non-uniform cell seeding in the final cytotoxicity calculation. The # of cells used is less than the cells needed for Release assays & Flow Cytometry. Flow cytometry & Release assays require a seeding density of 100K target cells increasing the number of effector cells to the millions. The visual observation of ADCC or CDC on the images can be convincing to conclude the functionality of antibodies or complements.

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A rapid 3D tumor spheroid analysis method using the Celigo Imaging Cytometry
Leo L. Chan, Scott Cribbes, Sarah Kessel, Olivier Dery, Catherine Swingler, Dmitry Kuksin, Tim Smith, Jean Qiu, Maria Vinci, Lisa Patterson, Sue Eccles

• Celigo Imaging Cytometer is a versatile and powerful tool for 3D tumorspheroid analysis
• Assays such as measuring optimal seeding density for forming tumorspheroids and viability measurement have been demonstrated
• Advanced assays such size measurement, growth inhibition, invasion into matrigel®, migration on to collagen and HUVEC, and tissue invasion that have been performed by manual microscope observation can now be easily quantified using the automated imaging cytometry method

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The Power Decoder simulator for the evaluation of pooled shRNA screen performance
Jesse Stombaugh, Abel Licon, Žaklina Strezoska, Joshua Stahl, Sarah Bael Anderson, Michael Banos, Anja van Brabant Smith, Amanda Birmingham, Annaleen Vermeulen

Power Decoder (written in R and Python) simulates shRNA pooled screening experiments in silico to allow for the estimation of a screen’s statistical power. Populations of shRNAs were engineered in such a way that the magnitude of depletion and enrichment was known, then using the negative binomial distribution, an in silico model was developed to successfully resemble data from an actual laboratory experiment.

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Scientific News
DNA Damage Seen in Patients Undergoing CT Scanning
Along with the burgeoning use of advanced medical imaging tests over the past decade have come rising public health concerns about possible links between low-dose radiation and cancer.
Yeast Cells Use Signaling Pathway to Modify Their Genomes
Researchers at the Babraham Institute and Cambridge Systems Biology Centre, University of Cambridge have shown that yeast can modify their genomes to take advantage of an excess of calories in the environment and attain optimal growth.
New Material Forges the Way for 'Stem Cell Factories'
Researchers have discovered the first fully synthetic substrate with potential to grow billions of stem cells. The researchcould forge the way for the creation of 'stem cell factories' - the mass production of human embryonic (pluripotent) stem cells.
New Measurements Reveal Differences Between Stem Cells for Treating Retinal Degeneration
By growing two types of stem cells in a “3-D culture” and measuring their ability to produce retinal cells, a team lead by St. Jude Children’s Research Hospital researchers has found one cell type to be better at producing retinal cells.
Researchers Identify Critical Genes Responsible for Brain Tumor Growth
After generating new brain tumor models scientists have identified the role of a family of genes underlying tumor growth in a wide spectrum of high grade brain tumors.
Growing Spinal Disc Tissue
Scientists develop new method for growing spinal disc tissue in the lab for combating chronic back pain.
A New Path Towards a Universal Flu Vaccine
New research suggests it may be possible to harness a previously unknown mechanism within the immune system to create more effective and efficient vaccines against this ever-mutating virus.
Potential New Class of Cancer Drugs
Scientists have found a way to stop cancer cell growth by targeting the Warburg Effect, a trait of cancer cell metabolism that scientists have been eager to exploit.
Human Trials of Manufactured Blood Within Two Years
The first human trials of lab-produced blood to help create better-matched blood for patients with complex blood conditions has been announced by NHS Blood and Transplant.
How Anthrax Spores Grow in Cultured Human Tissues
New findings to help predict risk and outcomes of anthrax attacks.
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