SuperScript® III CellsDirect™ cDNA Synthesis Kit

Catalog number: 18080300

Invitrogen™  Related applications: Reverse Transcription

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The SuperScript® III CellsDirect cDNA Synthesis Kit is optimized for synthesizing first-strand cDNA directly from a mammalian cell lysate without first isolating the RNA. Lysis and reverse transcription are performed in the same tube, and the resulting first-strand cDNA is ready to use in cloning and PCR. For real-time quantitative RT-PCR, see the note below. The kit has the following advantages:

• Compatible with a wide range of mammalian cell types grown under different treatment conditions
• Single-tube format minimizes reagent loss, sample loss, and handling time
• Total lysate volume is used in first-strand cDNA synthesis reaction, providing greater yields with a limited number of cells and allowing for detection of rare transcripts
• SuperScript® III Reverse Transcriptase, with reduced RNase H activity and higher thermal stability, produces high yields of cDNA in the first-strand synthesis reaction, for greater sensitivity and enhanced detection of rare transcripts
• Generates high-quality cDNA for use in a variety of applications, including cloning and PCR
• Simple protocol takes less than 2 hours

How it works
In traditional RT-PCR, RNA is first isolated from cells in a time-consuming procedure that can lead to a loss of material. Using the SuperScript® III CellsDirect cDNA Synthesis System, the cells are lysed and the cDNA is generated from the lysate in a single tube with minimal handling and no sample loss. DNase I is added to eliminate genomic DNA prior to first-strand synthesis. This kit has been optimized for small cell samples, ranging from 10,000 cells down to a single cell (as measured by serial dilution). The use of SuperScript® III Reverse Transcriptase ensures high specificity and high yields of cDNA from small amounts of starting material—as little as 10 pg total RNA. After synthesis, the first-strand cDNA can be amplified with specific primers by PCR without intermediate organic extractions or ethanol precipitations.

For Research Use Only. Not for use in diagnostic procedures.


Reverse Transcriptase: SuperScript® III
Downstream Application: Reverse Transcriptase PCR (RT-PCR), Real-Time Quantitative PCR (qPCR)
Reaction Format: Separate Components
Sample Type (General): Cells
Product Size: 100 reactions
Optimal Reaction Temperature: 50° C

Contents & storage

The SuperScript® III CellsDirect cDNA Synthesis Kit includes:
• Resuspension Buffer (1 ml )
• RNaseOUT™ Recombinant Ribonuclease Inhibitor (40 units/µl) 200µl
• DNase I (1 U/µl) 500 µl
• 10X DNase I Buffer 160 µl
• 25 mM EDTA 120 µl
• Oligo(dT)20 (50 µM) 120 µl
• 10 mM dNTP Mix 100 µl
• SuperScript™ III RT (200 units/µl) 100 µl
• 5X RT Buffer 600 µl
• 0.1 M DTT 100 µl
E. coli RNase H (2 U/µl) 100 µl
• HeLa Total RNA (10 ng/µl) 10 µl
• Forward Control Primer (10 µM) 10 µl
• Reverse Control Primer (10 µM) 10 µl
E. coli RNase H (2 U/µl) 100 µl
• HeLa Total RNA (10 ng/µl) 10 µl

Store all components at -20°C.


Manuals & protocols

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