I am having trouble with this quantification method. I am a CS major working in
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Question
I am having trouble with this quantification method. I am a CS major working in a developmental biology lab so I do apologize before hand if this question is dumb.
The MAK2 model for qPCR is: "Dn = (Dn-1) + (k)ln(1+((Dn-1)/k))", "Dn" is the value at cycle "n". "D0" and "k" are the only two adjustable parameters that determine "Dn". this is the paper: http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0012355.
How would I go about implementing this? As an example here is data from 30 cycles of qPCR: "610 618 627 632 628 634 634 628 634 642 637 643 653 666 684 717 787 923 1197 1716 2638 4077 5461 7007 8561 9994 11278 12382 13382 14252". The 5PSM and LinRegPCR quantification methods both yield an initial template fluorescence value of ~1. I think I am reading something wrong in the paper because I am just not seeing it haha. Thanks for your time, I am pretty sure I left something out so if you need more information let me know. -Sam
Explanation / Answer
I can help you find the efficiency (En) and the 'k' constant value. I did not find how to calculate Do. Also,
if this is not want you wanted to know then I'm sorry.
Dn=(Dn -1) + k ln [1+(Dn-1)/k]
where Dn= 14252, Dn-1=13382 . using these values you can calculate the k value from above equation.
also, En = Dn -[Dn-1]/Dn-1
= 14252-13382/14252
= 870/13382
En=0.065
however, if you know the background flurescence ( if the given fluorescence value~ 1is background flurescence (Fb)) then use this formula and calculate Fn first,
Fn= Dn+ Fb (Fn is fluorescense at n cycle)
then use Fn value obtained, to calculate Do, as follows: Fn = Do*En +Fb
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