Editing BioMicroCenter:RTPCR Protocol for Sample QC (section)

== Preparing Standards ==
 
This procedure creates seven standards ranging from ~20nM to 0.3nM with which to create the standard curve.  In preparing these standards 1:100 dilutions are created. After this step the standards are ready to use, they do not need to be diluted any further, and they can be kept at 4° for up to seven days.

* S1 – add 2uL of Phix control to 198uL of EB (1:100)

* S2 – add 50uL of S1 to 50uL of EB (1:200)

* S3 – add 50uL of S2 to 50uL of EB (1:400)

* S4 – add 50uL of S3 to 50uL of EB (1:800)

* S5 – add 50uL of S4 to 50uL of EB (1:1600)

* S6 – add 50uL of S5 to 50uL of EB (1:3200)

* S7 – add 50uL of S6 to 50uL of EB (1:6400)

Vortex '''well''' and spin down in between each dilution

<span style="color: purple">Illumina Phix concentrations vary from lot to lot.  It is recommended that you pool multiple tubes of Phix, separate them out into one time use aliquots (~2uL) and quantify one aliquot (we use the KAPA Library Quantification kit for qPCR). This will allow you to have consistent concentrations across multiple preparations of your standards.</span>