ExProfile™ Gene qPCR Arrays

Introduction

For high-throughput profiling of gene expression

The ExProfile™ Gene 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. In each 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. A cDNA pool, containing reverse transcript products from total RNA of 10 different tissues, was used as the qPCR validation template.

A universal real-time PCR condition was developed for easy profiling and analysis of  the gene expression in a high-throughput fashion. The All-in-One™ First-Strand cDNA Synthesis Kits and qPCR Mix Kits are the recommended and supported RT-PCR reagents for use with the ExProfile™ gene qPCR arrays. These reagents have been optimized to produce high sensitivity, efficiency, and specificity.

Key advantages

Validated mRNA primers

  • Each primer pair is designed using a proprietary algorithm and has been experimentally validated

Robust performance

  • Sensitive – Detects as low as 4 copies of RNA using ExProfile gene qPCR array and recommended reagents/conditions
  • Broad linearity – Simultaneously detects mRNAs at different expression levels
  • Reproducible – High reproducibility (R2> 0.99) for inter-array and intra-array replicates

Pre-arranged groups, or customized groups

  • Pre-arranged cancer-related groups
  • Pre-arranged pathway-related groups
  • Customized gene arrays for focused study

Gene qPCR array experimental work flow

Figure 1. Gene qPCR array experimental work flow.

 

Performance Data


Linear Range and Sensitivity (spike-in control RNA)
Broad linear range and high sensitivity

Figure 2. Broad linear range and high sensitivity

Mouse total RNA with serially diluted Spike-in control RNA were reverse-transcribed using All-in-One first strand cDNA synthesis kit. The reverse-transcribed cDNA samples were detected using All-in-One qPCR mix and spike-in control specific primers deposited in a 96-well plate. The resulting Ct values were plotted against the log5 of the amounts of spike-in control RNA. The data demonstrated a broad linear dynamic range from 4 to 1.6*106 copies of input RNA as well as high sensitivity.



Positive calls with a range of total RNA
High positive calls with as little as 15.36 ng of total RNA

Figure 3. High positive calls with as little as 15.36 ng of total RNA

Different amounts of MCF_7 total RNA (1000, 200, 40, 8, 1.6ng) were analyzed with the Human Breast Cancer Gene qPCR Array (PAG-HGBE96-01).The percentage of positive calls (Ct < 35) is plotted against the input amount of total RNA in each qPCR reaction.



Inter-Array Reproducibility
High inter-array reproducibility

Figure 4. High inter-array reproducibility


Two ExProfile™ qPCR gene array replicates (plate A and B) were analyzed using human total RNA (10-tissue mix) on the Bio-Rad iQ5. The Ct values of the replicate plates were plotted against each other. R2 > 0.99 was observed for high inter-array reproducibility. R2 > 0.99 was also observed for intra-array reproducibility (data not shown).