Complete and haplotype-specific sequence assembly of segmental duplication-mediated genome rearrangements using targeted CRISPR-targeted ultra-long read sequencing (CTLR-Seq)
Bo Zhou, GiWon Shin, Stephanie U. Greer, Lisanne Vervoort, Yiling Huang, Reenal Pattni, Marcus Ho, Wing H. Wong, Joris R. Vermeesch, Hanlee P. Ji, Alexander E. Urban
In this preprint the authors report on a method, CTLR-Seq, that combines Sage Science’s HLS-CATCH with low input Oxford Nanopore sequencing(using the MinION platform). The researchers are studying complex and highly repetitive genomic regions associated with neuropsychiatric disorders (mutations at 22q11.2 and at 16p11.2). Long read nanopore sequencing is used to resolve large segmental duplications, copy number variations, and large deletions.
Stanford University, Stanford CA
KU Leuven, Flanders Belgium
Ultra-low input single tube linked-read library method enables short-read second-generation sequencing systems to generate highly accurate and economical long-range sequencing information routinely generate highly accurate and economical long-range sequencing information
Zhoutao Chen, Long Pham, Tsai-Chin Wu, Guoya Mo, Yu Xia, Peter L Chang, Devin Porter, Tan Phan, Huu Che, Hao Tran, Vikas Bansal, Justin Shaffer, Pedro Belda-Ferre, Gregory Humphrey, Rob Knight, Pavel Pevzner, Son Pham, Yong Wang and Ming Lei
This study describes the TELL-Seq™ barcode linked read method developed by Universal Sequencing Technology Corp, and presents validation data for the process. TELL-Seq uses a transposase enzyme method that is linked to a bead (TELL-Bead) to insert an 18 bp molecular barcode into illumina sequencing libraries. The method allows economical analysis of haplotype phasing and structural variation in genomes or metagenomic analyses. The workflow requires low DNA input (0.1-5 ng) and is completed in a single tube.
Universal Sequencing Technology, Canton MA
Bioturing, Inc., San Diego CA
University of California, San Diego
CRISPR–Cas9/long-read sequencing approach to identify cryptic mutations in BRCA1 and other tumour suppressor genes
Tom Walsh, Silvia Casadei, Katherine M Munson, Mary Eng, Jessica B Mandell, Suleyman Gulsuner, Mary-Claire King
In this short report, researchers studying young-onset breast cancer used the SageHLS and HLS-CATCH method to purify high molecular weight targets containing the BRCA1 locus. Affected individuals in this study showed normal sequence based on gene panel and whole exome sequencing. Using long read (~10kb) and low input PacBio® HiFi™ sequencing on the HMW targets the researchers identified an SVA retrotransposon insertion in intronic regions of the BRCA1 locus that are homologous to a region on chr1. They were further able to show how BRCA1 transcription was affected.
University of Washington, Seattle WA
Journal of Medical Genetics
Prech Uapinyoying, Jeremy Goecks, Susan M. Knoblach, Karuna Panchapakesan, Carsten G. Bonnemann, Terence A. Partridge, Jyoti K. Jaiswal, and Eric P. Hoffman
Scientists researching muscular disorders present a method for studying gene expression of large RNA transcripts. The PacBio Iso-Seq method allows the sequencing of large transcripts for the identification of isoforms and alternative splicing. However, it has not been amenable to accurate quantitation of expression. Here the scientists combine Iso-Seq with PacBio’s HiFi high consensus accuracy method to show differential gene expression of large transcripts in muscles as well as identify and quantitate previously unannotated ones.
BluePippin was used to size select 5-10kb cDNA fragments for PacBio Iso-Seq library construction, and a second size selection step to remove unligated adapters.
Center for Genetic Medicine Research, Children’s Research Institute, Washington D.C.
Department of Genomics and Precision Medicine, The George Washington University School of Medicine and Health Sciences, Washington, D.C.
National Institute of Neurological Disorders and Stroke, NIH, Bethesda, MD
Computational Biology Program, Oregon Health and Science University, Portland, OR
Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, Binghamton University, Binghamton, NY
Fully phased sequence of a diploid human genome determined de novo from the DNA of a single individual
Ilya Soifer , Nicole L. Fong, Nelda Yi, Andrea T. Ireland, Irene Lam, Matthew Sooknah, Jonathan S. Paw, Paul Peluso, Gregory T. Concepcion, David Rank, Alex R. Hastie, 5 Vladimir Jojic, J. Graham Ruby, David Botstein, Margaret A. Roy
Researchers at Calico Life Sciences in a collaboration with the platform manufacturers PacBio (sequencing) and BioNano (optical mapping) produced a fully phased de novo assembly of a human genome. 10X Genomics linked-read technology was also used, and a new method is presented that uses Illumina sequencing of flow cytometry-sorted metaphase chmormosomes.
>20KB High-Pass was used on the BluePippin for the construction of SMRTbell library for PacBio RSII sequencing.
Calico Life Sciences LLC, South San Francisco,CA
Pacific Biosciences, Menlo Park, CA
Bionano Genomics, San Diego, CA
G3:Genes/Genom/Genetics (Early online)