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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
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.
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.
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.
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.