About QuestTaq™ qPCR PreMix
QuestTaq™ qPCR PreMix contains all reagents necessary to perform unbiased quantitative PCR with DNA having 5-methylcytosine (5mC), 5-hydroxymethylcytosine (5hmC), and glucosyl-5-hydroxymethylcytosine (g5hmC) composition. It features a unique DNA polymerase and optimized buffer system for amplification of DNA having modified/unmodified cytosines. It also includes an intense dsDNA-specific fluorescent dye (SYTO® 9) for sensitive/precise DNA detection. QuestTaq™ qPCR PreMix is ideal for locus-specific 5-hydroxymethylcytosine detection when used in conjunction with the Quest 5-hmC Detection Kit™ (Cat. Nos. D5410 & D5411 from Zymo Research) and the Quest 5-hmC Detection Kit™-Lite (D5415 & D5416).
|Equipment||Compatible with real-time PCR instruments that don’t require a passive reference dye (e.g., LightCycler® 480 (Roche), CFX96™ (Bio-Rad), etc.)|
|Storage||QuestTaq™ qPCR PreMix should be stored between -20 °C to -80 °C for maximum performance. Performance is guaranteed for 1 year from the time of receipt.|
|Unit Definition||One unit (U) enzyme of QuestTaq™ 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 (20 µl total volume) will contain 2 units of QuestTaq™ DNA polymerase.|
QuestTaq™ Polymerase yields the least cycle threshold variation (?Ct) versus the polymerases from Suppliers A, B, and C when amplifying DNA with modified cytosines. The figure shows ?Ct values from real-time PCR data generated using a LightCycler® 480 instrument. Equivalent amounts of template DNA having modified cytosines (5mC, 5hmC, and g5hmC) were compared to unmodified template (post-amplification) using QuestTaq™ qPCR PreMix or the polymerases from Suppliers A, B, and C.