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View additional product information for SuperScript™ IV VILO™ Master Mix - FAQs (11766050, 11756500, 11766500, 11756050)
14 product FAQs found
Yes, the TaqMan Cells-to-CT Express Kit includes Express ezDNase which can be added to the Express Lysis Solution for removal of gDNA during cell lysis. Express ezDNase is a double-strand specific, heat-labile DNase that will only digest gDNA, making it compatible with the downstream reverse transcription reaction. The enzyme is automatically inactivated during a heat kill step included in the reverse transcription program. To avoid the detection of gDNA, we recommend using a TaqMan Gene Expression Assay specifically designed to span an intron-exon boundary. Such assays are designated with an "_m1" suffix. More information can be found here.
Find additional tips, troubleshooting help, and resources within our Real-Time PCR and Digital PCR Applications Support Center.
The Invitrogen ezDNase Enzyme is a novel DNase that is highly specific for double-stranded DNA. It has no activity on single-stranded DNA in RT reactions (primers or probes), or on RNA. The enzyme is also thermolabile—it is inactivated quickly at temperatures typical for the SuperScript IV RT reaction (e.g., 50°C). The additional inactivation step is therefore not required in RT-qPCR applications.
Find additional tips, troubleshooting help, and resources within our PCR and cDNA Synthesis Support Center.
For RT-qPCR applications we recommend using the Invitrogen SuperScript IV VILO Master Mix (Cat. No. 11756050). The cDNA synthesis reaction setup with this master mix requires fewer pipetting steps and therefore reduces variation in the data. SuperScript IV RT, as a component of the master mix, offers the highest efficiency of cDNA synthesis step compared to competitors’ products.
Find additional tips, troubleshooting help, and resources within our PCR and cDNA Synthesis Support Center.
The only change is that the incubation time for the reverse transcription reaction has been reduced from 50 minutes to 10 minutes. All the other parameters and steps are the same.
Find additional tips, troubleshooting help, and resources within our PCR and cDNA Synthesis Support Center.
When compared with SuperScript III RT (and other manufacturers’ RTs) in a synthesis reaction for a 9 kb cDNA, SuperScript IV RT performed successful synthesis in just 10 minutes and did so with comparable (or improved) yield (as shown by gel band density).
Find additional tips, troubleshooting help, and resources within our PCR and cDNA Synthesis Support Center.
We have not created a one-step RT-qPCR kit containing SuperScript IV RT for real-time PCR. However, it is included in end-point one-step RT-PCR kits: https://thermofisher.com/ssiv-onestep
We also offer a two-step RT-qPCR kit containing SuperScript IV RT: https://www.thermofisher.com/vilo
For one-step RT-qPCR, we offer a kit with SuperScript III RT: https://www.thermofisher.com/order/catalog/product/117320200
The SuperScript IV VILO Master Mix contains SuperScript IV RT, a ribonuclease inhibitor, and a helper protein. The helper protein helps to increase the efficiency of the reverse transcription reaction and thus improve cDNA yield. This master mix formulation allows for a simpler reaction setup with less pipetting and less variation between samples.
The ezDNase enzyme is highly specific for double-stranded DNA and does not cause damage to RNA and single stranded DNA. The high specificity of its action makes the deactivation step largely irrelevant in the subsequent RT reactions. ezDNase enzyme can be inactivated at 55 degrees C incubation for 2 mins if needed.
For the 50 reaction SKUs, the no RT control can be for 50 reactions. For the 500 reaction SKUs, the no RT control can be for 250 reactions, which is way more than enough for control reaction purpose.
This is to help ensure that the biases from using either oligo (dT)18 or random hexamer primers alone will be eliminated, although the data indicate that using random hexamers alone provides quantitation just as good as the combination of the two primers. Some scientists want the combination because they feel it could be better for data quality.
SuperScript IV VILO Master Mix has increased thermostability, allowing the RT incubation temperature to be raised to deal with the structural complexity often associated with GC-rich templates. Our tests show that compared to other cDNA synthesis kits or master mixes, the raised reaction temperature leads to minimal loss of cDNA synthesis efficiency for SuperScript IV VILO Master Mix, suggesting that it is the best choice if high reaction temperature is needed for cDNA synthesis from GC-rich templates.
The recommended reaction temperature for SuperScript IV VILO Master Mix is 50 degrees C. R&D has tested the stability of SSIV VILO at higher temperatures and it works very well even at 65 degrees C.
SuperScript IV VILO Master Mix uses SuperScript IV Reverse Transcriptase (RT) in the optimized master mix formulation, whereas SuperScript VILO Master Mix and SuperScript VILO kits use SuperScript III Reverse Transcriptase. SuperScript IV RT has a significantly improved performance profile over SuperScript III RT in thermostability, sensitivity, yield, processivity, and the ability to synthesize cDNA efficiently from a wide variety of RNA samples, even those of suboptimal purity and integrity.
SuperScript IV VILO Master Mix is an upgrade from SuperScript VILO Master Mix and cDNA Synthesis Kit. It is designed to be used for cDNA synthesis in the two-step RT-qPCR applications for gene expression analysis.