Accuris qMAX Green One-Step RT-qPCR Kit

• RNA to cDNA to qPCR, in one tube
• High purity enzyme formulation for enhanced stability and performance
• Accuris Hot-Start Taq allows for preparation at room-temperature
• Blue dye facilitates pipetting and visualization in plates
• Available for green fluorescence (PR2120 Series) or probe detection (PR2121 Series

qMAX Green One Step kits incorporate our proprietary intercalating dye which exhibits higher fluorescent and lower PCR inhibition than other popular dyes such as SYBR. They are compatible with standard and fast cycling protocols and provide increased sensitivity, speed and reproducibility for a broad range of samples and targets. The polymerase mix is available with different levels of ROX reference dye for compatability with all qPCR instruments.

The qMAX Green One-Step RT qPCR Kit is a ready-to-use 2X master mix and companion thermostable reverse transcriptase for use in highly sensitive real-time RT-PCR assays in which intercalating dye-based detection provides the option of a post amplification melt profile.

The kit is powered by HS Taq DNA Polymerase, qMAX MMLV derived Reverse Transcriptase, and an optimized buffer chemistry for robust first-strand cDNA synthesis and real-time PCR in a single tube. The mix delivers earlier quantification cycle values (Ct) and broad range detection for increased sensitivity, speed, reliability and reproducibility.

The qMAX Green One-Step qPCR Mix can be used to quantify a specific target RNA from either total RNA or mRNA while reducing the number of pipetting steps and time to result.

Storage: Upon receipt, immediately store at -20°C. Avoid excessive freeze/thaw cycles. When stored as directed, this product will retain its activity for 12 months from date of receipt. The product may also be stored at 4°C for up to one month.

Limitations of Use: For research purposes only. Not intended for therapeutic or diagnostic use.

Quality Control: Accuris enzymes and reagents are tested under general assay conditions for activity, reproducibility, efficiency, heat activation, sensitivity, and absence of nuclease contamination and nuclease activity. This product is manufactured under a comprehensive quality management system, following ISO 9001:2008 standards.

General Guidelines:

• Reaction Mix: The reaction mix has been optimized to perform with maximum sensitivity and efficiency. The use of additives is not required or recommended.
• 20X Thermostable RTase: The 20X Thermostable Reverse Transcriptase is blended with a potent RNase Inhibitor.
• Template: Use 10pg to 100ng total RNA per reaction (or a minimum of 0.01pg mRNA per reaction).
• Reverse Transcription: Recommended incubation is 45°C for 10 minutes. For regions of high secondary structure, incubation temperatures up to 55°C may be used.
• Primers: Primers should have a predicted melting temperature of approximately 60°C, using default Primer 3 settings (http://frodo. wi.mit.edu/primer3/).
• Amplicon: : Optimal extension is achieved at 72°C. The optimal amplicon length should be 80bp to 200bp and should not exceed 400bp. When comparing qMAX Green One-Step RTqPCR Mix with a reagent from an alternative supplier, we strongly recommend amplifying from a 10-fold template dilution series. Loss of detection at low template concentration is the only direct measurement of sensitivity. An early Ct value is not an indication of good sensitivity, but rather an indication of reaction speed.

Reaction Setup: Thaw the qMax Green One-Step Kit and vortex briefly. Set up reaction as follows:

For other volumes, adjust the amount of each component accordingly.

Protocol: Program the qPCR instrument using the following conditions, acquiring data, on the appropriate channel:

• 1 cycle, 45°C for 10 minutes. For RNA with a high degree of secondary structure, use 55°C.
• 1 cycle, 95°C for 2 minutes for an initial denaturation and polymerase activation.
• Perform 30-40 cycles of :
  • 95°, 5 seconds (denaturation)
  • 60° - 65°C, 20-30 seconds (do not exceed 30 seconds and do not use temperatures below 60°C)

Packaging: