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Frederick National Laboratory for Cancer Research

6 ARTICLES PUBLISHED IN JoVE

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JoVE Journal

RNA Secondary Structure Prediction Using High-throughput SHAPE
Sabrina Lusvarghi *1, Joanna Sztuba-Solinska *1, Katarzyna J. Purzycka 1, Jason W. Rausch 1, Stuart F.J. Le Grice 1
1RT Biochemistry Section, HIV Drug Resistance Program, Frederick National Laboratory for Cancer Research

High-throughput selective 2' hydroxyl acylation analyzed by primer extension (SHAPE) utilizes a novel chemical probing technology, reverse transcription, capillary electrophoresis and secondary structure prediction software to determine the structures of RNAs from several hundred to several thousand nucleotides at single nucleotide resolution.

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Neuroscience

A High Content Imaging Assay for Identification of Botulinum Neurotoxin Inhibitors
Krishna P. Kota 1,6, Veronica Soloveva 2,6,7, Laura M. Wanner 3,6, Glenn Gomba 4,6, Erkan Kiris 3,5,6, Rekha G. Panchal 6, Christopher D. Kane 2,6,7, Sina Bavari 6
1Perkin Elmer Inc., 2Henry M. Jackson Foundation, 3The Geneva Foundation, 4ORISE, 5Frederick National Laboratory for Cancer Research, 6Division of Molecular and Translational Sciences, US Army Medical Research Institute of Infectious Diseases, 7DoD Biotechnology High Performance Computing Software Applications Institute (BHSAI), Telemedicine and Advanced Technology Research Center (TATRC), US Army Medical Research and Materiel Command (USAMRMC)

Botulinum neurotoxin is one of the most potent toxins among Category-A biothreat agents, yet a post-exposure therapeutic is not available. The high content imaging approach is a powerful methodology for identifying novel inhibitors as it enables multiparameter screening using biologically relevant motor neurons, the primary target of this toxin.

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Biology

SpOT the Correct Tissue Every Time in Multi-tissue Blocks
Anna Coffey 1,2, Michael D. Johnson 3, Deborah L. Berry 3
1Center for Advanced Preclinical Research (CAPR), Frederick National Laboratory for Cancer Research, 2Leidos Biomedical Research, Inc., 3Department of Oncology, Georgetown University

The purpose of the Specimen Orientation Tag (SpOT) is to function as an orientation tool to aid in individual tissue identification in multi-tissue paraffin blocks. These protocols demonstrate how it is constructed easily from common, low-cost histology materials and serves as a reliable visual marker in paraffin blocks and sections.

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Chemistry

Synthesis of Cd-free InP/ZnS Quantum Dots Suitable for Biomedical Applications
Matthew A. Ellis 1, Giovanna Grandinetti 2, Katye M. Fichter 1
1Department of Chemistry, Missouri State University, 2Center for Molecular Microscopy, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research

In this protocol, the synthesis of Cd-free InP/ZnS quantum dots (QDs) is detailed. InP-based QDs are gaining popularity due to the toxicity of Cd2+ ions that may be released through nanoparticle degradation. After synthesis, QDs are solubilized in water using an amphiphilic polymer for use in biomedical applications.

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Biochemistry

Fully Processed Recombinant KRAS4b: Isolating and Characterizing the Farnesylated and Methylated Protein
Constance Agamasu 1, Peter Frank 1, Shelley Perkins 1, Timothy Waybright 1, Simon Messing 1, William Gillette 1, Andrew G. Stephen 1
1NCI RAS Initiative, Cancer Research Technology Program, Frederick National Laboratory for Cancer Research

Prenylation is an important modification on peripheral membrane binding proteins. Insect cells can be manipulated to produce farnesylated and carboxymethylated KRAS4b in quantities that enable biophysical measurements of protein-protein and protein-lipid interactions

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Genetics

Optimization for Sequencing and Analysis of Degraded FFPE-RNA Samples
Yelena Levin *1, Keyur Talsania *1,2, Bao Tran 1, Jyoti Shetty 1, Yongmei Zhao 1,2, Monika Mehta 1
1NCI CCR Sequencing Facility, Frederick National Laboratory for Cancer Research, 2Advanced Biomedical and Computational Sciences, Frederick National Laboratory for Cancer Research

This method describes the steps to improve the quality and quantity of sequence data that can be obtained from formalin-fixed paraffin-embedded (FFPE) RNA samples. We describe the methodology to more accurately assess the quality of FFPE-RNA samples, prepare sequencing libraries, and analyze the data from FFPE-RNA samples.

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