Taq DNA polymerase, ready-to-use PCR and qPCR premixes, and reverse transcription kits for first-strand cDNA synthesis. Tell us your instrument and chemistry and we match the reagents to them.
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| Product | What it does | Use it for |
|---|---|---|
| Taq DNA polymerase | Thermostable polymerase that leaves a 3′ A overhang | Routine PCR, colony PCR, genotyping, TA cloning |
| PCR premixes | 2X mix of polymerase, buffer, Mg²⁺ and dNTPs | Fast, consistent end-point PCR setup |
| qPCR premixes | PCR premix with a fluorescent dye or formulated for probes | Real-time quantification of DNA or cDNA |
| Reverse transcription kits | Reverse transcriptase, primers and buffer for cDNA synthesis | Turning RNA into cDNA before PCR or qPCR |
Tell us the features you need, such as hot-start, a premix with gel loading dye, a specific ROX level or genomic DNA removal before reverse transcription, and we match the kit.
Dye or probe. Dye-based mixes (SYBR Green type) fluoresce when they bind any double-stranded DNA. They are economical and work with any primer pair, and a melt curve after the run confirms a single product. Probe mixes detect a sequence-specific hydrolysis probe, which adds specificity and allows multiplexing.
ROX. Some instruments need a passive reference dye to normalise well-to-well differences. Using the wrong level does not ruin a run, but it adds noise. Check your instrument:
| ROX level | Instruments (examples) |
|---|---|
| High ROX | Applied Biosystems StepOne and StepOnePlus, 7000, 7300, 7900HT |
| Low ROX | Applied Biosystems 7500, ViiA 7, QuantStudio series |
| No ROX needed | Bio-Rad CFX, Roche LightCycler, Qiagen Rotor-Gene |
The guide to choosing a qPCR master mix covers hot-start chemistry, fast cycling and efficiency checks in more detail.
In two-step RT-qPCR you make cDNA first, then use it in separate qPCR reactions. One cDNA stock can be tested for many genes and kept for later. In one-step RT-qPCR both reactions run in one tube with gene-specific primers, which saves handling and suits high-throughput testing of a few targets.
For two-step work, a mix of oligo(dT) and random primers gives even coverage of mRNA. RNA quality decides everything that follows, so start from a good extraction: the RNA extraction kits and the PCR vs qPCR vs RT-qPCR guide explain how the steps fit together.
| Component | Volume | Final |
|---|---|---|
| 2X qPCR premix | 10 µL | 1X |
| Forward primer (10 µM) | 0.4–0.8 µL | 200–400 nM |
| Reverse primer (10 µM) | 0.4–0.8 µL | 200–400 nM |
| Template DNA or cDNA | 1–2 µL | Up to about 100 ng gDNA |
| Nuclease-free water | To 20 µL |
A common three-step profile is an initial activation at 95 °C, then 40 cycles of 95 °C for 10–15 seconds and 60 °C for 30–60 seconds, followed by a melt curve. Activation times differ between hot-start chemistries, so use the times in the manual.
RT-PCR adds a reverse transcription step that copies RNA into cDNA before amplification. qPCR measures amplification in real time with a fluorescent dye or probe. Combined, RT-qPCR quantifies RNA. Some people also use RT-PCR to mean real-time PCR, so check which is meant.
It depends on the instrument. Many Applied Biosystems instruments use ROX as a passive reference, at a high or low level depending on the model. Bio-Rad CFX, Roche LightCycler and Rotor-Gene instruments do not need it. Tell us your instrument and we match the mix.
No. A qPCR reaction needs a fluorescent dye or probe chemistry and a mix optimised for real-time detection. Some end-point PCR premixes also contain loading dyes that interfere with fluorescence readings.
Oligo(dT) primers copy polyadenylated mRNA from its 3′ end. Random primers copy all RNA, including partly degraded and non-polyadenylated RNA. Gene-specific primers give the highest sensitivity for one target and are standard in one-step RT-qPCR. A mix of oligo(dT) and random primers is a good default for two-step work.
At -20 °C. Enzyme mixes in glycerol do not freeze solid and can be used straight from the freezer, kept on ice while in use. Protect dye-containing qPCR premixes from light and avoid repeated freeze-thaw of primers and templates by aliquoting.
qPCR / RT-PCR
End-point PCR tells you whether a sequence is there, qPCR tells you how much, and RT-qPCR does the same for RNA. Here is what changes between them and the reagent list for each.
4 minute read
qPCR / RT-PCR
A 2X qPCR master mix looks like a commodity until it does not match your instrument or chemistry. This guide covers dye vs probe, ROX levels, hot-start, setup volumes and how to validate efficiency.
4 minute read
qPCR / RT-PCR
Your controls usually say what went wrong. A positive control, a no-template control, a no-RT control and one dilution of the sample separate template, inhibition and primer problems.
3 minute read
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