The Genomics Shared Resource provides an integrated set of tools and services for high throughput technologies for applied genomics. The Microarray division is funded through the NCI CCSG, RPCI, SUNY Buffalo and chargeback revenues. cDNA and genomic BAC clones, such as those identified in the Genomics division projects, are offered as an arrayed and archived resource. Investigators can request microarray services including RNA/DNA isolation, Cy3 and Cy5 probe generation, hybridizations, custom-made oligonucleotide and genomic arrays, Agilent, Affymetrix and Illumina arrays, scanning of microarrayed slides, image analysis, normalization of data and web-based interface with an Oracle database. The Genomics division is funded through grants, contracts, and chargeback revenues, and contributes to the application of human and mouse BAC clones for downstream functional analysis. It provides the infrastructure for automation through robotics for large-scale projects. In addition to the cDNA libraries, seven BAC/PAC genomic libraries are available for clone selection, characterization, mapping, and distribution. The Genotyping division is funded through the NCI CCSG, RPCI support and chargeback revenues. Investigators can request large scale SNP detection, SNP analysis, loss of heterozygosity and copy number determination, quantitative gene expression and methylation studies using the Sequenom MassARRAY and Illumina GoldenGate and Infinium systems.

Scheduling

Investigators should contact Jeff Conroy (716-845-3361) for details. Priority is given to NCI-funded investigators with RPCI affiliation, and as determined by the RPCI Genomics Shared Resource Steering Committee. 

User Fees

There is a chargeback system for users of the RPCI Genomics Shared Resource. The fees vary depending on the affiliation of the investigator, cost of materials and the extent of services requested from the user (see fee schedule).  Billing is done at the completion of the service.

Material Transfer Agreements

GENOMIC SERVICES ONLY: Roswell Park Cancer Institute must receive a completed and signed Material Transfer Agreement (Academic MTA, Commercial MTA) from both Academic and Industrial users before RPCI BAC and /or PAC clones can be distributed. Any questions with this agreement should be directed to Dr. Richard Matner, RPCI Tech Transfer (716-845-8192).

Policies

Project Description - Each RPCI investigator is required to submit a brief description of their project, and need for microarray, genotyping, methylation or genomic services. Please include program affiliation and grant funding (NCI, etc.). Projects may be reviewed by the Steering Committee before services are performed.

Agilent, Affymetrix, Illumina and BAC copy number, LOH and gene expression microarrays

Microarray Hybridization- All microarray hybridization requests must be ordered on line: https://rpcilims.roswellpark.org.

RNA - 0.5 - 1 µg total RNA is required per Agilent and Illumina gene expression or miRNA array. Note: It is recommended to treat RNA samples with DNase I. All RNA samples must have a A260/280 between 1.8 - 2.0, A260/230 > 1.6 and a bioanalyzer RIN > 7.0. It is required that the user provide the concentration, volume, buffer and preparation method of the sample. Degraded RNA samples can be analyzed (i.e. paraffin, formalin fixed tissues) with Illumina DASL gene expression and miRNA arrays.

DNA - 1 µg high molecular weight genomic DNA is required per BAC, Illumina or Affymetrix array hybridization. It is required that the user provide the concentration, A260/A280 ratio and the volume of the sample. Low molecular weight DNA samples can be analyzed (i.e. paraffin, formalin fixed tissues), but results may be more difficult to interpret.  The facility cannot guarantee aCGH results from low molecular weight DNA samples.

Tissues - A minimum of 10⁶ cells or 25mg of tissue is required for RNA/DNA preparation. All samples must be fresh or snap frozen in liquid nitrogen and stored at -80°C.  FFPE samples can be used with BAC arrays (DNA - copy number) and the Illumina DASL gene expression and miRNA arrays (RNA).

Data Analysis - The facility provides basic image analysis using platform dependent software (ImaGene - BAC arrays, GenomeStudio - Illumina, Feature extraction & GeneSpring - Agilent).  For BAC arrays, a plot will be generated along with a spreadsheet detailing LogR ratios of all normalized spots. This spreadsheet along with supporting image files will be available for data analysis by investigators. The GSR staff cannot be expected to clearly understand the subtleties of each investigators's experiments. Therefore, it is up to each investigator to interact with RPCI's bioinformatics staff to complete the final data analysis. Our approach analyzes each experimental parameter to ensure high data quality. Some of the parameters measured are analyzed the same way for many different kinds of experiments. Others, especially time trial studies, depend strongly on the experiments being done and the decisions from the experiments that can only be made by the investigator.

Microarray Printing - Investigators can select individual oligos, BACs or cDNA clones from the archived clones sets and have them arrayed as custom slides. Investigators are responsible for selecting the oligo-clone name/plate IDs from the database and uploading them in the on line ordering system. Investigators can also provide oligos, cDNA clones or cDNA PCR products for custom arrays. PCR products must be received in 384-well Genetix plates (X6004), 20µl at 500ng/µl in 20% DMSO.  All BAC & cDNA clones must be received in 96-well plates as frozen glycerol stocks and be certified phage free. Please include primers needed for PCR amplification of the inserts and growing requirements of the clones.

Genomic Services

Genomic Clone Requests - BAC and PAC clones can be ordered online from the facility. Please select the format you wish to receive the clones (DNA, glycerol, stab).  Roswell Park Cancer Institute must receive a completed and signed Material Transfer Agreement (Academic MTA, Commercial MTA) from both Academic and Industrial users before RPCI BAC and /or PAC clones can be distributed. Any questions with this agreement should be directed to Dr. Richard Matner, RPCI Tech Transfer (716-845-8192).

Genotyping and Methylation Services - Sequenom and Illumina

Genotyping - All genotyping requests must be received with a completed, signed requisition. Information for SNP sequences to be used for iPLEX oligo and OPA design must be emailed in spreadsheet format consistent with the example described below. DNA samples to be genotyped must pass quality control measurements and also have a corresponding spreadsheet provided.

DNA - It is required that the user provide the concentration, A260/A280 ratio and the volume of the sample.  Low molecular weight DNA samples can be analyzed (i.e. paraffin, formalin fixed tissues), but results may be more difficult to interpret and should be discussed before submitting.  Sequenom: 100-200ng genomic DNA is required for SNP genoyping and 1ug for methylation analysis.    Each microgram of DNA for methylation analysis will be bisulfite treated, resulting in enough tempate for 50 reactions.  Illumina: 250ng (via picogreen quantitation) genomic DNA is required for targeted GoldenGate SNP genoyping and 500ng (via picogreen quantitation) for Infinium whole genome genotyping.

SNP Sequence - Simply submit a target SNP sequence(s) for any genome and we will design and manufacture a custom SNP Assay for you. To design custom multiplexes for the MassARRAY and Illumina GoldenGate Systems, you can use SNPs from a variety of sources, including SNPs you already have and those from public databases. 

Methylation Sequence - Sequenom: Simply submit a target meth sequence(s) and we will design and manufacture a methylation assay for you. It is recommended that you provide at least 1,500 bp of sequence and that the region of interest be indicated in the sequence file. Sequemom EpiTyper requires amplicons from 200-600bp, so some CpG regions may have to be assayed using several PCR primers.  Illumina: The human methylation 27 array contains >27,000 CpG sites for whole genome analysis.  No sequence submission required.