About ZymoTaq™ qPCR Premix
The ZymoTaq™ qPCR PreMix product contains all the reagents needed to perform quantitative PCR and other molecular downstream analysis such as high-resolution melt (HRM) analysis of DNA methylation and other general real-time PCR assays. It features a hot-start DNA polymerase and buffer system optimized for the amplification of bisulfite-treated DNA. It also includes an intense dsDNA-specific fluorescent dye, SYTO 9®, for sensitive real-time DNA quantification. This technology can also be found in the Femto Quantification Kits (Cat. No. E2005, E2006, E2007) for low copy detection of DNA from human, bacteria, or fungi.
|Storage||Store at -20°C for up to 12 months. Minimize exposure to light. Avoid repeated freeze/thawing of reagents.|
|Unit Definition||One unit (U) enzyme of ZymoTaq™ DNA Polymerase is defined as the amount of enzyme required for the incorporation of 10 nmol dNTPs into an acid-insoluble form in 30 minutes at 72°C.|
|Enzyme Concentration||Reaction conditions at 1X (50 µl total volume) will contain 2 U of ZymoTaq™ DNA polymerase.|
Real-Time Amplification Plot. This figure shows robust real-time amplification of 10 ng of bisulfite-converted human DNA using the ZymoTaq™ qPCR PreMix and primers designed to amplify a region of the MGMT promoter.
Reliable Standards for the Quantification of Human DNA: Human DNA Standards performed in duplicates 10-fold serial dilutions ranging from 20 ng to 20 fg.
Suggested Reaction Setup (50 µl):
Note: The final concentration of MgCl2 in the reaction (above) is 2.75 mM. If required, scale reaction reagent volumes accordingly to optimize the MgCl2, primer, and/or template concentrations.
Suggested Conditions for qPCR:
Set the real-time PCR instrument to excitation and emission wavelengths of 465 nm and 510 nm, respectively.
Note: A setting that is compatible with SYBRgreen® should be compatible with the SYTO 9® dye in this kit.
*Note: Add 15 to 30 seconds to the extension time for each additional kb over 1 kb. Make adjustments to the temperature if necessary.