CCR Genomics Core lists over 1000 registered members in its iLab management and scheduling software with 325 research groups of which 267 are from NCI and the remaining 58 from 13 other NIH institutes. These institutes include NIAID, NHGRI, NEI, NINDS, NHLBI, NIAMS, NIDDK, NIMH, NIA, NIAAA, NIDCD, NINCR, and NINR. Of the 52 labs/branches listed under CCR, 35 utilize our services. Those labs include: Basic Research Laboratory, Cancer Inflammation Program, Developmental Therapeutics Branch, Endocrine Oncology Branch, Experimental Immunology Branch, Experimental Transplantation and Immunology Branch, Genetics Branch, HIV and AIDs Malignancy Branch, HIV Dynamics and Replication, Laboratory of Biochemistry and Molecular Biology, Laboratory of Cancer Biology and Genetics, Laboratory of Cell and Developmental Signaling, Laboratory of Cell Biology, Laboratory of Cellular and Molecular Biology, Laboratory of Cellular Oncology, Laboratory of Genitourinary Cancer Pathogenesis, Laboratory of Genome Integrity, Laboratory of Human Carcinogenesis, Laboratory of Immune Cell Biology, Laboratory of Metabolism, Laboratory of Molecular Biology, Laboratory of Protein Dynamics and Signaling, Laboratory of Receptor Biology and Gene Expression, Laboratory of Timor Immunology and Biology, Lymphoid Malignancies Branch, Neuro-Oncology Branch, Radiation Biology Branch, Radiation Oncology Branch, Surgery Branch, Thoracic and Gastrointestinal Oncology Branch, Thoracic and Oncologic-Surgery Branch, Vaccine Branch, Women's Malignancies Branch, Center of Advanced Preclinical Research, and NCI Rare Tumor Initiative Program. Available Core services include: Sanger Sequencing is carried out using one 96 capillary 3730XL and two newly acquired 3500xL Genetic Analyzers, replacing two obsolete ABI 16-capillary 3130XLs. These upgraded instruments have increased read-length 800 base pairs, turn-around time with higher number of samples per run, improved data precision and maintains a 20% smaller footprint. Data are analyzed and made available within one business day. The current charges are $2.00 for electrophoresis only and $7.00 for the sequencing reactions, clean-up and electrophoresis. The Core also has a variety of bulk plate cost options including sets of 16 and full 96 well trays. Illumina Next Generation Sequencing is performed with two different platforms, the MiSeq Desktop Sequencer, NextSeq 500 and NextSeq 550 which was recently added to the Core. Researchers can submit pre-made libraries or purified DNA/RNA for library construction and subsequent sequencing. The Core will also perform quality control (QC) on all libraries prior to sequencing. Individuals meet with Core members to discuss the best experimental designs and costs are offset by subsidies provided by the Office of Science and Technology Resources (OSTR). NanoString Digital Gene Expression Analysis System is capable of direct quantification of individual mRNAs in a biological sample without the use of enzymes or amplification. NanoString designs each CodeSet and then investigators bring their samples to the core to be processed. The cost of the Codesets is subsidized by the OSTR for NCI, CCR investigators. Core charges range from $5/sample to $24/sample for NCI staff depending on what services and supplies are necessary. The cost for Non-NCI staff is $48/sample. Bio-Rad QX200 Droplet Digital PCR System provides absolute quantification of target DNA, including measurement of viral loads and microbial quantification; detection of copy number variations and rare mutations; absolute quantification of mRNA and miRNA and quantification of NGS libraries. Typically, after discussion with Core staff and if necessary BioRad for assay development, customers bring prepared samples in a sealed plate ready to be applied to Automatic Droplet Generator. Results are usually available within 24 h. Data analysis software is available for use on customers' own computer. Current costs $5/sample for NCI and non-NCI customers. Fluidigm C1 Single Cell Autoprep System for genomic analysis of single cells is an automated single-cell isolation and preparation system that provides an easy and reproducible workflow to process 96 single cells for genomic analysis including RNA-Seq, Whole Genome Amplification and sequencing and Gene Expression Profiling on Biomark HD qPCR. Customers contact the Core manager to schedule training and access to the instrument. The core also provides access to the Nexcelom Vision CBA image cytometry system to determine cell size, number and viability prior to C1single cell isolation as well as Keyence All-in One microscope for annotation. Subsidies for this technology are provided by the OSTR. The instrument is also made available as a part of the single cell repertoire through the CCR Single Cell Analysis Facility. Agilent TapeStation 2200 & 4200 Systems allow for reproducible and user-independent quality control of DNA and RNA samples and libraries. QC services are provided to customers obtaining NGS services from our Core as well as the CCR Sequencing Facility at the Frederick National Laboratory. The Core charges range from $4/sample to $5/sample for NCI staff depending on number of samples and what supplies are necessary. The new 4200 system allows for fully automated sample processing for up to 96 samples with results being obtained in as little as 1-2 minutes per sample, or less than 90 minutes for 96 samples while requiring only 1-2 uL of sample per run. In addition, the CCR Genomics Core functions as a support lab, containing instruments for various applications for use by CCR investigators. The Core has several robotic liquid handlers to include the Agilent Bravo and the Formulatrix Mantis. These robotic devices support our NGS workflow but also are made available for robotizing assay development and performance. Other available equipment for investigators include Qubit, Sage Science PippinHT and the Keyence BZ-X700 microscope. Analytical software for the various technologies is also made available. So far in FY18, the Core has managed over 3370 iLab requests and processed 21,629 samples for Sanger sequencing, 612 for Nanostring, 8925 for ddPCR and 6188 for tapestation. The Core has been involved in 191 different NGS projects featuring the MiSeq and NextSeq 500/550 platforms. Partnerships with NIDCD and Dr. Jing Huang in NCI continue with single cell isolation using the Fluidigm C1 with the 96 well IFC. The Big Dye Program where the Core purchases Big Dye in bulk discounted quantities and then aliquots it and sells it to Core users at cost continues to thrive and save the NCI money. Thus far in in FY18 we have had 37 requests totaling $26,496 in sales. The program is available for all investigators within CCR, even if they do not use the core for sequencing. Other Core achievements have included acknowledgement as well as Core members included as collaborators and authors on manuscripts when extensive collaborative efforts have been involved (Nepal et al., Hepatology, 2018; Lissa et al., 2018; Peretti et al., Cell Host Microbe, 2018; Lu et al., Mol. Ther, 2018; Keen et al., Genome Announc., 2017). Liz Conner, Core Manger, served as the primary site visit reviewer of the Genomics Core at the National Institutes of Child Health and Development in May of 2018. Dr. Conner also serves as the NCI Chair of the Occupational Safety and Health Committee.
