To determine lysis efficiency, RT-qPCR should be evaluated across a 5-log dilution of cells. Resuspend cells in Cell Resuspension Solution and perform lysis of 10 – 100,000 cells per 50 µl reaction. Perform one-step RT-qPCR with the cell lysates and check the RT-qPCR efficiency using a gene of interest (target/primers should be validated using pure RNA first). The ideal efficiency should be 90%-110%. Efficiency <90% often indicates RNA damage. Efficiency >110% with delayed initial Cq may imply potential inhibitors present in the RT-qPCR.
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