The mission of the La Jolla Institute for Allergy &Immunology (LIAI) is the study of immune cells in health and disease. Their areas of interest cover an extremely wide spectrum of projects including: (i) analysis of allele- specific chromatin state and DNA methylation and hydroxymethylation status in asthma, embryonic stem cells and cancer;(ii) changes in chromosome architecture during the differentiation of cytolytic T lymphocytes;(iii) defining allele-specific global transcriptional output from a variety of immune cells obtained from patients with diseases such as asthma, allergy, sickle cell disease, cancer, cardiovascular disease, diabetes and various autoimmune diseases, as well as from relevant animal models of human disease. Next-generation sequencing is one of the newest and most powerful tools to understand these processes. The study of genomes, epigenomes and transcriptomes provides unbiased information on how genes function and interact with one another in response to outside environmental signals and in the chromatin context to maintain normal physiology or to cause disease. The expected surge of interest in this technology nationwide has inter- fered with the access of NIH-funded LIAI investigators to local and even national facilities for next-generation sequencing, forcing them to pay high overhead costs and wait many weeks for their samples to be sequenced. Although we have a SOLiD 5500xL sequencer, this is already being used to capacity and moreover is not suited for certain applications needed for our funded NIH projects, including genome-wide DNA methylation and hydroxymethylation by bisulfite sequencing analysis;relating disease-associated single-nucleotide poly- morphisms to changes in chromatin structure or gene expression in cis;mapping nucleosome positioning genome-wide;analysis of long-range DNA looping by Hi-throughput chromosome capture (Hi-C);and obtaining high-quality sequence information from very small numbers of sorted immune cell populations. We are therefore requesting funds for purchase of an Illumina HiSeq 2500, a unique, high-throughput, cost effective next-generation sequencing system with highly sensitive Time Delayed Integration (TDI) line scanning and four CCD sensors for dual surface imaging to sequence massively in parallel up to 55 gigabases (Gb) per day. It uses a straightforward sequencing-by-synthesis method with reversible terminator chemistry, the most desirable and versatile sequencing configuration, ideal for a wide range of genetic analyses and functional genomics applications. If awarded, this versatile instrument will be available to all scientists at LIAI following an in-house next-generation sequencing training course, and will undoubtedly allow us to make timely and efficient progress on our NIH-supported projects. The SOLiD sequencing system presents challenges in achieving long sequencing read lengths and studying DNA methylation status;the Ion PGM sequencer can produce longer reads but through-put is much lower.

National Institute of Health (NIH)
Office of The Director, National Institutes of Health (OD)
Biomedical Research Support Shared Instrumentation Grants (S10)
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Special Emphasis Panel (ZRG1-GGG-A (30))
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Birken, Steven
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La Jolla Institute
La Jolla
United States
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