Posted by Jing Zhang on February 25, 2008 11:59 PM|Permalink
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Here are some questions for Jing and the rest of the class:
1. Assuming that, the S/N~10 for 10 copies of template (see figure 4), what is the limit of detection for the fluorescence detector in this device? Express the LOD as 3xStd. dev. of the background and assume that the analyte is FAM. You may need to make other assumptions.
2. How did the authors prove (or could prove) that the error in peak areas is originating from the gel interface?
Assume that the fluorescence intensity is linear with the DNA concentration and after 30 complete PCR cycle, the amount of DNA still grow in the exponential way, the LOD should be 3 copies of DNA. But I am not sure when the DNA copy number goes too low, the first few cycles are efficient.
The author didn't prove that the error in peak areas is originating from the gel interface. I am not sure how to prove it since there is another variable: the injection efficiency.
For the second question, I am still not sure why the author uses peak area to look at the microdevice uniformity here,instead of peak height. Since the initial number of pUC19 and PCR cycle numbers are the same, it is supposed to get the same namount of DNA at the end of the PCR. Will the gel interface cause peak height to be different with each other?. If it will, why do we use peak area instead of peak height? I found other places in this paper use peak height more.
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Comments
Here are some questions for Jing and the rest of the class:
1. Assuming that, the S/N~10 for 10 copies of template (see figure 4), what is the limit of detection for the fluorescence detector in this device? Express the LOD as 3xStd. dev. of the background and assume that the analyte is FAM. You may need to make other assumptions.
2. How did the authors prove (or could prove) that the error in peak areas is originating from the gel interface?
Posted by: Edgar Arriaga | February 26, 2008 05:39 PM
Assume that the fluorescence intensity is linear with the DNA concentration and after 30 complete PCR cycle, the amount of DNA still grow in the exponential way, the LOD should be 3 copies of DNA. But I am not sure when the DNA copy number goes too low, the first few cycles are efficient.
The author didn't prove that the error in peak areas is originating from the gel interface. I am not sure how to prove it since there is another variable: the injection efficiency.
Posted by: Jing Zhang | February 27, 2008 01:44 AM
For the second question, I am still not sure why the author uses peak area to look at the microdevice uniformity here,instead of peak height. Since the initial number of pUC19 and PCR cycle numbers are the same, it is supposed to get the same namount of DNA at the end of the PCR. Will the gel interface cause peak height to be different with each other?. If it will, why do we use peak area instead of peak height? I found other places in this paper use peak height more.
Posted by: Yixiao Sheng | February 28, 2008 06:29 AM