Sample Preparation and Hybridization Protocols

SpliceExpress™ microarrays for alternative splicing support a variety of standard protocols, reagant kits and equipment. Total RNA is the most common starting point for experiments with SpliceExpress™. For full service customers, we usually request 5µg of total RNA per sample per array. This is used for RNA purification and quality testing as well as for hybridization. The actual hybridization can be done with less than 1µg of RNA.

Several commercially available amplification and labeling kits work well. Kits that amplify RNA without 3′ bias are preferable because splicing may occur near the 5′ end of the gene. Jivan probes are optimized for use with standard hybridization protocols. We can make more detailed recommendations when an order is placed.

Scanners from Axon and Agilent can read the arrays and extract features.

For data analysis, Jivan's splice arrays are compatible with standard software packages such as SpotFire, GeneSpring, Partek and Resolver. Full service customers can also benefit from Jivan's proprietary data analysis methods, including deconvolution of exon-exon junction and exon-intron junction signals to identify splice products that span multiple probes.

For validation by rt-PCR, Jivan has a database of PCR primer designs capable of detecting the splice variants for each SpliceExpress™ product.