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The qPCR assay facilitates calculation of the optimal number of Endpoint PCR cycles to use for final library amplification. The assay described below (using the PCR Add-on and Reamplification Kit; Cat. No. 208) is applicable for use with the following library preparation methods:
All CORALL RNA-Seq V2 with UDI library prep kits (Cat. No. 171 – 186-186, 219-22, 233-234),
All QuantSeq V2 with UDI library library prep kit (Cat. No. 191 – 196, 222-223, 225),
QuantSeq-Pool (Cat. No. 139),
LUTHOR HD (Cat. No. 204, 221).
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,
LUTHOR HD Pool (Cat. No. 205).
Additional Reagents Required
The qPCR assay to determine optimal PCR cycle numbers for library amplification is performed using an aliquot of purified double-stranded cDNAcDNAs, prior to the final library amplification.
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*For QuantSeq-Pool, if you have >7 pools, the PCR Add-on and Reamplification Kit V2 is required for qPCR. If you have <6 pools, the provided kit solutions for Library Amplification (PM, P5, P7, PE) are sufficient for qPCR and final amplification.
ATTENTION! The use of SYBR Green I-containing qPCR mastermixes from other vendors is not recommended.
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