The qPCR assay requires SYBR Green I dye (final concentration of 0.1x in the reaction mix) and a real-time PCR machine. The TeloPrime Full-Length cDNA Amplification V2 Kits (Cat. No. 013) contain sufficient reagents to perform a qPCR and endpoint PCR for each sample. Lexogen also offers a TeloPrime PCR Add-on and Reamplification V2 Kit (Cat. noNo. 018) ,for 16 reactions if additional PCRs are required.
Determine the cycle number at which the fluorescence reaches at 80% of the maximum. If using the same volume of cDNA for the endpoint PCR, this is the cycle number to be used for the endpoint PCR.
If more cDNA is used for the endpoint PCR than for the qPCR, the cycle number may need to be adjusted. For example, if 2 µl of cDNA is used for the qPCR assay, and 9 µl will be used for endpoint PCR, subtract 2 cycles to account for using ~4x more cDNA as template for the final cDNA amplification. See Appendix B of the TeloPrime V2 User Guide for more details.