Making Light Work of Differential Display: Genomyx Corporation Introduces fluoroDD Kit

Faster, cheaper, cleaner. All of these words describe Genomyx's latest addition to its line of products for differential display analysis of gene expression. The new Fluorescent Differential Display Adaptor Kit (fluoroDD Kit) provides fluorescently labeled primers that can be used with Genomyx's HIEROGLYPHTM mRNA Profile Kits to allow for a rapid and nontoxic alternative to radioactive labeling of differential display products. Fluorescent Differential Display with fluoroDDTM mRNA Profile System. Paired samples produced from human fetal and adult liver were run after priming with different anchored and arbitrary primers. ARP = arbitrary primers, TMR-AP = TMR-labeled anchored primer, HFL = human fetal liver, HAL = human adult liver While differential display analysis was embraced by many looking for a fast way to identify and isolate differentially expressed gene sequences, numerous technical difficulties hamper its usefulness. Frequently encountered problems include the generation of irreproducible fragment patterns, the inability to resolve fragments adequately, and inaccurate or inefficient isolation of interesting bands. Genomyx Corporation first introduced an integrated reagent and instrument system which addressed some of these tough spots and enhanced the potential of differential display. The genomyxLR® DNA Analysis System, which contains a long-bed gel unit (61 cm x 33 cm), power pack, and dryer, improves resolution in both the large and small fragment range. The HIEROGLYPH mRNA Profile Kits, with their unique anchored and arbitrary primers, provide expanded coverage of expressed sequences, while simultaneously streamlining downstream characterization analysis. The HIEROGLYPH technology is based on a primer design allowing high annealing stringencies to maximize reproducibility. Each of the five kits comes with all 12 oligo(dT) anchoring primers (each with a different two base combination at the 3'end) and a unique set of four different arbitrary primers (from the twenty available). The 12 oligo(dT) anchored primers (31-mers) each include the T7 R NA polymerase promoter sequence at the 5' end. The twenty possible arbitrary primers are 26 nucleotides long and each contains a 10 nucleotide core with M13 reverse sequencing primer at the 5'end. Following identification and recovery of putative differentially expressed bands, the resulting cDNA fragments, upon reamplification with M13 reverse and T7 promoter primers, are designed to go directly into a cycle sequencing reaction, eliminating a timeconsuming cloning step. The newest products in the Genomyx line are the fluoroDD Adaptor Kit, the genomyxSC Fluorescent Imaging Scanner, and the Band Excision Workstation, all of which work in concert to allow researchers to convert their Differential Display Gene Expression studies from radioactivity to fluorescent detection. At the core of these products is the fluoroDD Adaptor Kit which builds upon the success of the HIEROGLYPH mRNA Profile System by providing a set of TMR-labeled anchored primers which are designed to work with the HIEROGLYPH Kits. The fluoroDD kit significantly reduces the amount of time it takes to go from RNA to differential display pattern by eliminating the need for overnight exposures. In addition, because of the stability of the label, batch processing of samples is possible, decreasing the time required for sample preparation and thereby increasing throughput. By labeling on one strand only, less complex band patterns are generated, improving reproducibility and recovery (Y. Tokuyama and J. Takeda, Biotechniques, 18:424-425, 1995) of bands of interest. A unique feature of the Genomyx fluorescent approach to DD is the ability to locate and easily cut out the candidate differentially expressed cDNA fragments from the fluoroDD gel. This capability is provided by the Excision Workstation, consisting of a gel platform, a coordinate grid of vertical and horizontal lines (the physical grid), and an excision bridge. Genomyx has also developed the Virtual Grid, a digitized image of the physical grid, which can be overlaid via software on the fluorescent gel image so that the exact location of potential bands of interest can be determined. Following excision, the dried gel can be immediately rescanned to confirm accurate band recovery. For more information, contact Genomyx at 800-943-6663 or info@genomyx.com or visit the company web site at www.genomyx.com.

Making Light Work of Differential Display: Genomyx Corporation Introduces fluoroDD Kit

The Scientist Staff

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