序列检测引物

Most pre-designed TaqMan assays are supplied as a 20x assay concentrate containing 18 uM of each primer and 5 uM of probe which provides a 900 nM of each primer and 250 nM of the probe (for gene expression assays) and 200 nM of each probe (for SNP genotyping assays) at the final 1x concentration.

In order to make a 20x working stock from custom oligos, the oligos need to be diluted as follows: To make a 20x Assay stock for Gene Expression assays, make 100 uM stocks of both forward and reverse primers, and TaqMan probe. Add 18 ul of each primer and 5 ul of the TaqMan probe, which gives 41 ul volume. Add 59 ul of 1X TE to bring the volume to 100 ul of 20X TaqMan Gene Expression Assay. To make a 20x Assay stock for a Genotyping Assay, make 100 uM stocks of both forward and reverse primers, and both TaqMan probes. Add 18 ul of each primer and 4 ul of each TaqMan probe, which gives 44 ul volume. Add 56 ul of 1X TE to bring the volume to 100 ul of 20X TaqMan SNP Genotyping Assays.

The main difference between Custom Standard DNA Oligos and Sequence Detection Primer (Cat. Nos. 4304970, 4304971, 4304972) is that Sequence Detection Primers come in the guaranteed specific amount and concentration that is ordered. Whereas Custom DNA Oligos can be selected by the Synthesis Scale. The scale of synthesis (25 nmol, 50 nmol, 200 nmol, 1 µmol, or 10 µmol) refers to the starting scale and not the guaranteed final amount.

You can find more information about the Synthesis Scale by going to the Custom Standard DNA Oligos page and clicking on the question mark next to “Synthesis Scale.”

In addition, the sequence detection primers are desalted, whereas the custom DNA oligos can be purified by HPLC, PAGE, and Cartridge.

Find additional tips, troubleshooting help, and resources within our PCR and cDNA Synthesis Support Center.

We do not have pre-designed forward and reverse primers for GAPDH to be used in a SYBR Green dye-based real-time PCR assay. You can order either Sequence Detection Primers or Custom Standard DNA Oligos.

Find additional tips, troubleshooting help, and resources within our Real-Time PCR and Digital PCR Applications Support Center