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==Data Analysis== <ol> <li>Normalization</li> <ol type="A"> <li>CT values were obtained from the qPCR curves via the Biorad qPCR results</li> <li>CT values were subtracted from either the NTC CT value or positive control CT value (for quantitation)</li> <li>Cycle differences were averaged when technical replicates existed; e.g. for all experimental samples</li></ol> <li>Quantitation and Efficiency</li> <ol type="A"> <li>Positive control values were calculated from IDT dilutions (2 uL of 10 nM = 0.02 pmol) or Qubit measurements</li> <li>Qubit measurements (product)</li> <ol type="a"> <li>3100, 3210, and 2810 ng/mL</li> <li>Divide by 100X (Qubit measured sample was 100X qPCR template)</li> <li>Convert to pmol in 2 uL: 1.81*10^-3, 1.87*10^-3, 1.64*10^-3 pmol</li> <li>Average: '''1.8*10-3 pmol'''</li></ol> <li>Amount in the sample is 2^(cycle difference poscon-sample) * 1.8*10^-3 pmol</li> <li>From [[Daniel:Notebook/ComboLock/2017-2-13|bead math]], ~3.2 pmol of template should bind to bead, which means 3.2 pmol of C probe may bind and 3.2 pmol of ligation product may form, the theoretical maximum assuming 100% bead binding, 100% C probe binding, 100% wash efficiency, and 100% ligation efficiency</li> </ol></ol>
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