Comparison of RNA isolation and library preparation methods for small RNA sequencing of canine biofluids
Candice P. Chu, Mary B. Nabity
From the Department of Veterinary Pathobiology at Texas A&M, this study evaluates best practices for small RNA sequencing from canine biofluid samples (serum and urine). The study compares RNA-seq results after using three commercial RNA extraction kits for each biofluid type, and two sequencing library construction kits. A recommendation is provided for isolation/construction for each biofluid, with supporting data.
The PippinHT was used to isolate the small RNA libraries prior to sequencing.
Veterinary Clinical Pathology
Rory Bowden, Robert W. Davies, Andreas Heger, Alistair T. Pagnamenta, Mariateresa de Cesare, Laura E. Oikkonen, Duncan Parkes, Colin Freema, Fatima Dhalla, Smita Y. Patel, Niko Popitsch, Camilla L.C. Ip, Hannah E. Roberts, Silvia Salatino, Helen Lockstone, Gerton Lunter, Jenny C. Taylor, David Buck, Michael A. Simpson, Peter Donnelly
The potential of using Oxford Nanopore Technologies MinION sequencer is evaluated for Whole Genome Sequencing (WGS). A reference sample (NA112878) was sequenced alongside an individual with clinical indications. The authors applied novel bioinformatic approaches to the analysis and identify innovations that could significantly improve the platform’s potential for clinical WGS.
BluePippin was used to enrich DNA fragments >6kb using the High-Pass protocol prior to library construction.
Wellcome Centre for Human Genetics, Oxford UK
Hospital for Sick Children, Toronto CA
Oxford University, Oxford UK
National Institute for Health Research, Oxford UK
Children’s Cancer Research Institute, Vienna Austria
Transposon insertion profiling by sequencing (TIPseq) for mapping LINE-1 insertions in the human genome
Jared P. Steranka, Zuojian Tang, Mark Grivainis, Cheng Ran Lisa Huang, Lindsay M. Payer, Fernanda O. R. Rego, Thiago Luiz Araujo Miller, Pedro A. F. Galante, Sitharam Ramaswami, Adriana Heguy, David Fenyö, Jef D. Boeke & Kathleen H. Burns
A new technique is described; Transposon Insertion Profiling (TIPseq). The method uses vectorette PCR and paired-end Illumina sequencing to map (LINE-1, L1) retrotransposon insertions in the human genome. A detailed protocol is provided.
Pippin Prep is used to remove fragments below 400 bp from libraries prior to sequencing.
Johns Hopkins University School of Medicine, Baltimore, MD
NYU Langone Health, New York, NY,
Hospital Sírio-Libanês, São Paulo, Brazil
Universidade de São Paul, São Paulo, Brazil
Identification of whole blood mRNA and microRNA biomarkers of tissue damage and immune function resulting from amphetamine exposure or heat stroke in adult male rats
Luısa Camacho, Camila S. Silva, Joseph P. Hanig, Robert P. Schleimer, Nysia I. George, John F. Bowyer
A report from the USDA and others evaluate the effect of amphetamine and methamphetamine toxicity on immune systems and tissues using rat models. In particular, hyperthermic effects, due to the drug exposure or environmentally induces were studied. With hyperthermia, an increase in organ related transcripts were indicated and as was possible liver damage. In addition, changes to the immune system suggest modified risk of for HIV and other infections.
The PippinHT was used to isolated small RNA (15-35bp) Illumina libraries prior to sequencing.
Andrea Minio, Mélanie Massonnet, Rosa Figueroa-Balderas, Amanda M. Vondras, Barbara Blanco-Ulate, and Dario Cantu
Researchers used PacBio’s Iso-Seq method to sequence full-length cDNA transcripts from ripening Cabernet Savignon berries resulting in 1,5000 cultivar-specific genes. The authors discuss using transcriptome profiles for comprehensive referencing as an alternative to costly genome referencing.
The BluePippn was used to size select 1-2 kbp, 2-3 kbp, 3-6 kbp, and 5-10 kbp fractions for cDNA SMRTbell library construction for the Iso-Seq method.
University of California, Davis
G3: Genes, Genomes, Genetics