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View additional product information for Platinum™ SuperFi II PCR Master Mixes - FAQs (12368010, 12369010, 12368250, 12369250, 12368050, 12369050)
11 product FAQs found
Typically, for a 50 µL PCR reaction, we recommend using 25µµL of the 2X master mix so that its final concentration in the reaction is 1X.
Find additional tips, troubleshooting help, and resources within our PCR and cDNA Synthesis Support Center.
The dyes in the Platinum SuperFi II Green PCR Master Mix can interfere with the fluorescent readings on both Qubit fluorometer and Tapestation. To accurately quantify your samples, you should first use a PCR purification kit, such as the GeneJet PCR Purification Kit (Cat. No. K0701), to clean up your samples. Alternatively, you can use the Platinum SuperFi II PCR Master Mix (Cat. No. 12368010) which does not contain the interfering dyes, thus eliminating the need for a purification step.
Find additional tips, troubleshooting help, and resources within our PCR and cDNA Synthesis Support Center.
Good lab practices are important for long fragment amplification. These include using high-quality templates (pure, fresh, and intact) and fresh primer solutions. Reducing primer concentration to 0.2 µM may also improve the results.
Yes, Platinum SuperFi II DNA Polymerase works at both universal 60 degrees C annealing temperature and at annealing temperatures calculated with a Tm calculator.
Platinum SuperFi II DNA Polymerase produces blunt-ended PCR products that can be cloned directly into blunt-ended cloning vectors. TA cloning is also possible if 3' dA-overhangs are added after PCR. We recommend purifying the PCR products of Platinum SuperFi DNA Polymerase before adding the overhangs. The procedure for adding 3' dA-overhangs (TA cloning) includes the following steps:
- Purify the PCR product (e.g., with a PCR purification kit or phenol extraction/DNA precipitation). Before adding the overhangs, PCR product must be purified, as the strong proofreading activity of any remaining Platinum SuperFi DNA Polymerase will degrade the added 3' dA-overhangs.
- Perform 3' dA addition with a Taq DNA polymerase.
Reaction components:
Purified PCR product
0.2 mM dATP
1x Taq Buffer
1 U Taq DNA Polymerase
Incubate the reaction for 20 min at 72 degrees C.
- Proceed to TA cloning. For optimal ligation efficiency, we recommend using fresh PCR products, since 3' dA-overhangs can be lost during storage
Platinum SuperFi II reaction buffer is compatible with restriction digestion directly after PCR. Note that Platinum SuperFi II DNA Polymerase is not inactivated during PCR, thus purification of PCR products before restriction digestion is recommended if intact 5'- or 3'- overhangs are needed for cloning.
The green tracking dye (which consists of a blue and a yellow dye) is compatible with downstream applications such as fluorescent automated DNA sequencing, ligation, and restriction digestion. For applications that require analysis of PCR products by absorbance or fluorescence excitation, we recommend using colorless versions of the products (Platinum SuperFi II DNA Polymerase or Platinum SuperFi II PCR Master Mix) or purifying the PCR products prior to analysis.
The green tracking dye (which consists of a blue and a yellow dye) does not interfere with electrophoresis in E-Gel agarose gels. The sample should be diluted 2- to 20-fold for optimal separation using E-Gel agarose gels.
Platinum SuperFi II DNA Polymerase cannot read dUTP-derivatives or dITP in DNA templates, so the use of these analogues is not recommended. Platinum SuperFi II DNA Polymerase can incorporate 7-deaza-dGTP and radiolabeled dNTPs.
Platinum SuperFi II DNA Polymerase enables amplification of both AT-rich and GC-rich targets. Lower extension temperature (68 degrees C or even 65 degrees C) can be helpful for extremely AT-rich targets.
Platinum SuperFi II DNA Polymerase can amplify targets with high GC content (up to 75% GC) without any additional DNA melting agents. In cases of extremely GC-rich targets (>75% GC), addition of DMSO to final concentration of 5% is recommended.