Power SYBR™ Green Cells-to-CT™ Kit
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<i>Power</i> SYBR&trade; Green Cells-to-C<sub>T</sub>&trade; Kit
Invitrogen™

Power SYBR™ Green Cells-to-CT™ Kit

Green features
Ambion™ Power SYBR™ Green Cells-to-CT™ Kit is a fast, easy, and robust method to go directly from cultured cells toRead more
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Catalog NumberNo. of Reactions
440295340 reactions
4402955400 reactions
4402954100 reactions
Catalog number 4402953
Price (USD)
766.00
Each
Add to cart
No. of Reactions:
40 reactions
Price (USD)
766.00
Each
Add to cart

Ambion™ Power SYBR™ Green Cells-to-CT™ Kit is a fast, easy, and robust method to go directly from cultured cells to real-time PCR results. A breakthrough cell lysis and RNA stabilization technology eliminates the laborious and time-consuming RNA purification process completely. The kit includes sufficient reagents for 400 reactions. Power SYBR™ Green Cells-to-CT™ Kit is Powered by AmpliTaq Gold™ DNA Polymerase, LD, which provides the highest levels of specificity with standard real-time PCR.

• Includes optimized reagents to work efficiently, right out of the box
• Simple, effective Cells-to-CT™ technology enables sample preparation at room temperature in only 7 minutes, including DNase treatment
• Validated accuracy, exceptional reproducibility, and maximal sensitivity from 10 to 100,000 cells per sample; results equivalent to those from purified RNA

Extraordinary Value
The kit includes reagents for sample preparation integrated into a complete, optimized gene expression workflow including reverse transcription reagents and a high-performance PCR master mix with SYBR™ Green.

Extraordinary Ease
The protocol begins with a simple 7-minute sample preparation procedure. First, cells are washed in PBS, and then lysed for 5 minutes at room temperature; DNase treatment can be performed concurrently. The lysis reaction is stopped, and the sample is then ready for reverse transcription or storage at –20°C for up to 5 months. Because samples are processed directly in culture plates (96- or 384-well), sample handling and the potential for sample loss or transfer error are minimized, resulting in higher reproducibility. Following sample preparation, a portion of the cell lysate is added to an RT reaction followed by real-time PCR using the included master mix.

Extraordinary Performance
To demonstrate the performance of the kit, replicate experiments were run using either traditional RNA purification and real-time PCR analysis, or the Power SYBR™ Green Cells-to-CT™ Kit. The kit shows equivalent performance to results obtained using purified RNA for analysis of 10 to 100,000 cells. In addition, the sensitivity, efficiency, and dynamic range were shown to be superior to competitor lysate kits. Samples prepared with the kit have been tested for compatibility on various real-time PCR platforms including those from Applied Biosystems, Roche, and Bio-Rad.

Accessory Product:
The SYBR™ Green Cells-to-CT™ Control Kit (Cat. No. 4402959) contains a Xeno™ RNA control and primer set and is recommended to monitor the efficiency of cell lysis and amplification inhibition in Cells-to-CT™ reactions.

For Research Use Only. Not for use in diagnostic procedures.
Specifications
Detection MethodSYBR
PolymeraseAmpliTaq Gold DNA Polymerase
Reaction SpeedStandard
For Use With (Equipment)7000 System, 7200 System, 7300 System, 7500 System, 7700 System, 7900HT System,Applied Biosystems StepOnePlus™ Fast Real-Time PCR System, GeneAmp 5700, GeneAmp 9600, StepOnePlus™, Standard Mode, QuantStudio 6 and 7 Pro systems, QuantStudio 7 Flex system, QuantStudio 3 and 5 systems, QuantStudio 12k Flex system, QuantStudio 6 Flex system
Green FeaturesFewer resources used and less waste, Less hazardous
Label or DyeSYBR Green
Passive Reference DyeROX (Pre-mixed)
Product LineCells-to-CT™, SYBR™
Purification Time5 min.
Purity or Quality GradeLD (Low DNA)
Reverse TranscriptaseM-MLV
Shipping ConditionDry Ice
For Use With (Application)RT-PCR
Concentration20X
Hot StartBuilt-In Hot Start
No. of Reactions40 reactions
Sample TypeCells
Unit SizeEach
Contents & Storage
Store Stop Solution, DNase I and 20X RT Enzyme Mix at -20°C. Store Lysis Solution, 2X SYBR™ RT Buffer, and PowerSYBR™ Green PCR Master Mix at 4°C. Store Nuclease-free Water at room temperature.

Frequently asked questions (FAQs)

I'm seeing PCR products in the minus-RT control after performing my Cells-to-CT experiment. What does this mean?

If PCR products are seen in the minus-RT control reaction, but not in the no-template control, it indicates that genomic DNA remains in the sample and that genomic DNA was amplified in real-time PCR. Please follow the suggestions below:

- Ensure the DNase I is mixed thoroughly into the Lysis Solution.
- Use fewer cells per lysis reaction.
- Lyse cells using Lysis Solution that is at room temperature, and make sure that the lysis reaction occurs at room temperature.
You can also try increasing the incubation time of the lysis reaction to 8 minutes and/or using Lysis Solution that has been warmed up to 25 degrees C for cell lysis.

I'm getting PCR products in the no-template PCR control when performing a Cells-to-CT experiment. What could cause this?

PCR products in the no-template PCR control indicate that the sample is contaminated with DNA. More stringent steps need to be taken to control contamination.

I'm getting no PCR product or unexpected PCR products after performing a Cells-to-CT experiment. What could be the cause of this?

Please review the following possibilities and suggestions:

- A problem with adding or mixing the Stop Solution: ensure that the Stop Solution was added directly to the lysate, as components of the Lysis Solution may inhibit RT-PCR if not fully inactivated.
- The RNA was degraded: keep cells in PBS on ice before starting the cell lysis procedure.
- RNase in the sample was not completely inactivated: Too many cells could have been used or too much PBS left on the cells, diluting the lysis solution.
- The lysates sat too long before going to room temperature: Do not allow lysates to sit longer than 20 minutes at room temperature once the Stop Solution has been added.
- The sample does not contain the target RNA: Verify that the procedure is working by using the XenoRNA Control in the sample. Also check that your PCR primers can amplify your target under the PCR conditions you are using.

I ran out of stop solution for my Cells-to-CT experiment. Can I purchase it separately?

Yes, it is available in 1 mL aliquots (Cat. No. 4402960).

I have genomic DNA contamination in my Cells-to-CT reaction. How do I get rid of it?

1. Ensure that all medium is removed from the wells.
2. Wash with an equal volume of room temperature 1X PBS after the medium is removed.
3. Ensure that the reaction happens at room temperature (the lysis reaction may not reach room temperature if the plate is on ice, if the plate was quickly moved to the bench, or if a cold lysis solution was added).
4. Warm lysis solution to room temperature before adding to cells.
5. Allow the lysis reaction to proceed for 8 minutes at 25 degrees C.