RT-PCR (qPCR) Analysis

The qPCR Arrays are designed for profiling the expressions of pre-made or customized sets of coding-genes in various tissues or cells. The resulting differential expressions of profiled genes help researchers to identify those that are biologically significant and relevant to their research. An array can contain hundreds of primers for human genes or miRNA carefully chosen based on a thorough literature search of peer-reviewed publications. 

In a typical 96-well plate there are up to 84 pairs of qPCR primers and 12 wells of controls which are used to monitor the efficiency of the entire experimental process- from reverse transcription to qPCR reaction. Each pair of primers used in the qPCR arrays has been experimentally validated to yield a single dissociation curve peak and to generate a single amplification of the correct size for the targeted mRNA.

From the RNA/miRNA provided, first-strand cDNA is synthesized through reverse transcription which is later mixed with the qPCR reaction mix and distributed into the wells of the array and followed by the qPCR cycling. The qPCR result is analyzed with a Data Analysis System using the ΔΔCT method to perform fold-change analysis or simple statistical analysis of the expression level (CT or CP values) for each gene.

• Validated mRNA or miRNA primers: each primer pair is designed using a proprietary algorithm and has been experimentally validated. Primers validated for human, mouse, and rat are available- but primers for other species can be designed
• Robust performance
• Sensitive: detects as low as 4 copies of RNA; miRNA from as little as 10 pg of small RNA; or 20 pg of total RNA
• Broad linearity: simultaneously detects mRNAs at different expression levels
• Reproducible: high reproducibility for inter-array and intra-array replicates
• Pre-arranged (cancer or pathway-related groups) or customized groups for focused study
• Flexible assay design: arrays are available in 96-well and 384-well plates, with different layouts to choose form depending on the number of genes and replicates to be analyzed

Generally, for each 96 well plate, 0.02-1 ug total RNA is needed (low-abundance RNA may not be detected when using less than 10 ng total RNA), For miRNA arrays, 0.25-0.5 ug of small RNA or 0.5-1ug total RNA is needed for one 96-well plate. 

High quality cDNA is a prerequisite for accurate detection of gene expression; therefore RNA with A260/280 greater than 1.8 and free of genomic DNA or other residual contamination should be submitted.

For other formats or sample types please inquired for more details at info@picoimmune.com

Nucleic Acid Extraction and RNA Analysis Services are also available.