3 articles published in JoVE
RNA Secondary Structure Prediction Using High-throughput SHAPE Sabrina Lusvarghi*1, Joanna Sztuba-Solinska*1, Katarzyna J. Purzycka1, Jason W. Rausch1, Stuart F.J. Le Grice1 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.
Isolation of Immune Cells from Primary Tumors Stephanie K. Watkins1, Ziqiang Zhu1, Keith E. Watkins2, Arthur A. Hurwitz1 1Tumor Immunity and Tolerance Section, Laboratory of Molecular Immunoregulation, Cancer and Inflammation Program, National Cancer Institute - Frederick, 2KEWB Productions In this report, we describe a protocol for isolating highly purified populations of leukocytes that infiltrate tumors. This protocol is adapted from the Miltenyi Biotech protocol to enhance yield and purity for isolating cells from complex tumor tissue.
Amplifying and Quantifying HIV-1 RNA in HIV Infected Individuals with Viral Loads Below the Limit of Detection by Standard Clinical Assays Helene Mens1, Mary Kearney1, Ann Wiegand1, Jonathan Spindler1, Frank Maldarelli1, John W. Mellors2, John M. Coffin3 1The virology Core at the HIV Drug Resistance Program, NCI-Frederick, 2Division of Infectious Diseases, University of Pittsburgh, 3Department of Molecular Biology and Microbiology, Tuffts University Quantifying levels of HIV-1 RNA in plasma and sequencing single HIV-1 genomes from individuals with viral loads below the limit of detection (50-75 copies/ml) is difficult. Here we describe how to extract and quantify plasma viral RNA using a real time PCR assay that reliably measures HIV-1 RNA down to 0.3 copies/ml and how to amplify viral genomes by single genome sequencing, from samples with very low viral loads.