QuantiGene 2.0 Plex Assay Service

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QuantiGene at DNA Services Facility

The DNA Services (DNAS) facility at the University of Illinois at Chicago (UIC) provides a multiplex gene expression service using Panomics designed QuantiGene Plex 2.0 Reagent System. This system is designed to quantitate RNA (mRNA or lncRNA) molecules directly from cell lysates, blood (including whole blood, PAXgene blood, and dried blood spots), fresh and frozen tissue homogenates, FFPE tissue homogenates, total or mRNA enriched RNA, or in vitro transcribed RNA preparations. Multiplex assays are custom designed and can vary from 3 to 80 gene targets. A number of pre-designed gene array sets (including 27-30 plex endogenous control assays and 80-plex pathway sets) are offered. We also offer DNA CNV assays for 3-33 genes. Previously created genes are listed at www.panomics.com.

The advantages of this method include: (1) Direct detection of target RNA without reverse-transcription or complete RNA purification, (2) Effective on a wide variety of sample types, including FFPE tissue, even H&E stained slides (3) Heavily multiplexed assay allows for increased throughput (up to 96 samples/day), (4) All assays are performed/analyzed on the exact same sample aliquot, and (5) Rapid data acquisition. The downside of the approach, relative to quantitative PCR, is a smaller dynamic detection range (~3 orders of magnitude; a minimum of 2,000 transcripts per well are required for quantification). This approach is recommended for medium-to-large sample sets with medium-to-large number of target genes (e.g. >24 samples with >10 gene targets). These assays are NOT specific to animals, and can be developed for plants and microorganisms. A schematic of the assay is shown below.


The first step to utilizing the QuantiGene service as the DNAS Facility at UIC is to contact our staff – either Vaiva or Stefan – to discuss the approach and to determine if this approach is suitable for your project. Second, you will submit a list of mRNA or lncRNA sequences (or Genbank accession numbers are required when available) for assay development. If assays have previously developed, there is no additional fee, but first-time development fees are charged for novel assays. There are currently more than 5,500 genes in their catalog. Once your assay has been ordered (the DNAS facility will place orders for all necessary reagents, if requested), and you will need to prepare your samples for analysis. Sample preparation is performed in your laboratory using a lysis kit purchased from Affymetrix. Sample preparation is simple, and we can assist you in learning to perform this procedure. Samples are lysed and can be brought or shipped (on dry ice) to the DNAS facility. Please contact Vaiva before bringing or shipping samples. Upon receipt of the samples and the assay kit from Affymetrix, we begin analysis of your samples, as shown in the workflow below.

Before we run your entire plate, we will discuss with you the importance of determining the appropriate dilution factor for detection of all genes in your assay. Since some endogenous controls are strongly expressed, assays targeting these genes can be saturated in lysates that are not diluted. We highly recommend performing a test of your samples with endogenous control panels to (a) determine the appropriate endogenous controls for your study (there are panels available for human, mouse, and rat), and (b) determine the appropriate dilution to target your endogenous controls and your genes of interest without having to assess all samples at multiple dilutions. We further recommend including multiple endogenous control assays in your QuantiGene assay with different levels of expression (i.e. weakly expressing, moderately expressing, strongly expressing). Common endogenous control genes – such as ACTB or GAPDH – are strongly expressed, can be easily saturated, and do not always show the lowest coefficient of variation across datasets.

For more information and sample submission guidelines [1]


Please contact Vaiva Liakaite (vaiva@uic.edu/312-413-9290) or Dr. Stefan Green (greenDNA@uic.edu/312-996-7509) with any questions or inquiries.


PRICING:

General information about QuantiGene pricing is in the brochure, however, the final cost of the project depends on what kind of project you have. Please email us with your requests and we will individually discuss your own project and its costs.

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