The CellAmp Whole Transcriptome Amplification Kit (Real Time), Ver. 2 enables one to go from cells to cDNA and then use the generated cDNA as a template for real-time PCR. When using conventional techniques for working with trace amounts of nucleic acids, loss of product during purification steps is a frequent problem. However, this cells to cDNA kit ensures good product yield because loss-inducing purification steps for RNA and cDNA are not required.
The procedure begins with cell lysis, followed by a reverse transcription reaction with the dT adapter primer (RT dT Primer 2) to synthesize cDNA from mRNA. Next, terminal deoxynucleotidyl transferase (TdT) is used to add a dA tail to the synthesized cDNA. The resulting 3' dA tail containing cDNA product is used as direct template for PCR, resulting in fewer required amplification cycles.
In addition to moving from cells to cDNA, this product can be used to amplify cDNA from trace amounts of RNA.
The CellAmp Whole Transcriptome Amplification Kit (Real Time), Ver. 2 uses an additional exonuclease I treatment step, which helps suppress nonspecific primer-derived amplification and increases yields of low expression level genes.