Invitrogen™ TaqMan™ Gene Expression Cells-to-CT™ Kit

Description

• The TaqMan™ Gene Expression Cells-to-C T ™ Kit makes it possible to perform expression analysis directly from cultured cells without RNA purification
• This kit saves time and offers a simple workflow that is suitable for a few samples or can be easily incorporated into automated, high-throughput applications
• The TaqMan™ Gene Expression Cells-to-C T Kit is: • Complete —optimized workflow includes cell lysis reagents with gDNA removal, RT enzyme mix, buffer, and new TaqMan™ Gene Expression Master Mix • Fast —7-minute sample prep, including DNase treatment, at room temperature • Easy —lyse samples in a tube or directly in culture plates • Robust —perform gene expression analysis on 10–100,000 cells per sample; results equivalent to those from purified RNA • Efficient —contains sufficient reagents to generate 500 real-time PCR results from 100 starting samples Featuring a unique method for lysing cultured cells while removing genomic DNA and preserving RNA integrity, the TaqMan™ Gene Expression Cells-to-C T ™ Kit contains reverse transcription (RT) reagents for cDNA synthesis and TaqMan™ Gene Expression Master Mix for real-time PCR analysis
• TaqMan™ primer⁄probe sets are sold separately
• Simple 7-minute sample preparation: part of a complete workflow Whether you are using plates or tubes, the TaqMan™ Gene Expression Cells-to-C T ™ Kit, which uses the simple 7-minute sample preparation procedure outlined (see figure), is designed for 10–100,000 cultured cells⁄sample
• Cells are washed in PBS and lysed in solution for 5 minutes at room temperature; DNase treatment can be performed simultaneously
• Lysis is terminated at room temperature by a 2-minute incubation with Stop Solution
• The lysates are now ready for reverse transcription or storage at –20°C
• Because samples can be processed directly in culture wells (96 or 384 wells), sample handling and the potential for sample loss or transfer error are minimized, facilitating rapid, high-throughput processing
• Unlike old-fashioned, multi-step RNA isolation protocols, no heating, washing, or centrifugation steps are required
• The kit greatly simplifies a laborious 30–60 minute process and reduces it to 7 minutes
• The TaqMan™ Gene Expression Cells-to-C T ™ Kit workflow enables unsurpassed gene expression evaluation with any of the >700,000 TaqMan™ Gene Expression Assays
• This kit has been extensively tested for specificity with a broad selection of TaqMan™ Gene Expression Assays and shows performance equivalent to that obtained with purified RNA (see figure)
• Achieve unsurpassed performance and sensitivity Unlike some competitor kits that limit the amount of lysate in the RT reaction to 5%, the TaqMan™ Gene Expression Cells-to-C T ™ Kit can accommodate 45% of the total RT reaction volume as cell lysate
• Additionally, cDNA can comprise up to 45% of the real-time PCR reaction volume
• The large lysate volume in the optimized RT reaction, along with the large cDNA volume in the subsequent real-time PCR using the TaqMan™ Gene Expression Master Mix, lead to maximum sensitivity
• The master mix amplifies the target precisely and accurately, enabling the detection of small quantities of target, such as transcripts expressed at low levels
• The ability to detect limited target quantities was tested by mixing a constant number of human cells with various numbers of mouse cells
• The cell mixtures were prepared using the TaqMan™ Gene Expression Cells-to-C T ™ and assayed for a mouse-specific and human-specific gene
• Comparative data were generated in the same manner with RNA purified by traditional methodology
• The data show that RNA from as few as 10 mouse cells was detectable in a background of RNA from 10,000 human cells (see figure)
• The ability of the TaqMan™ Gene Expression Cells-to-C T ™ Kit to detect relative low abundance transcripts was equivalent to that of purified RNA; at low levels, it was superior
• Additionally, the performance of the TaqMan™ Gene Expression Cells-to-C T ™ Kit was compared to competitor lysate kits and to purified RNA
• Inputs of 100–100,000 cells⁄lysis reaction were examined
• The sensitivity of the TaqMan™ Cells-to-C T ™ Kit protocol was equivalent to that obtained with purified RNA, and it surpassed competitor lysates (see figure)
• Furthermore, the lack of inhibition at high cellular inputs and the low variation among technical replicates demonstrate the reliability of this approach for gene expression studies using cultured cells
• Proven performance, proven together All components of the TaqMan™ Gene Expression Cells-to-C T ™ Kit have been optimized for consistent and reliable performance
• This removes the guesswork involved in assembling separate sample preparation, RT, and real-time PCR kits
• And the TaqMan™ Gene Expression Cells-to-C T ™ Kit has been validated with TaqMan™ Gene Expression Assays for added quality assurance
• Accessory product The TaqMan™ Cells-to-C T ™ Control Kit was designed for use with the TaqMan™ Gene Expression Cells-to-C T ™ Kit
• This kit contains an endogenous control (ACTB) to normalize for sample loading variability and an artifical XenoRNA™ control and corresponding TaqMan™ Gene Expression Assay to monitor the effects of PCR inhibition
• Order Info Shipping Condition: Dry ice