ChIP DNA Quantitation with Low-Range PicoGreen Standard Curve
Hello everyone,
We’ve been having problems using the PicoGreen assay to determine our final ChIP DNA samples prior to sample prep for use with the ILMN system. While the high-range standard curve is reasonably linear, the lower portion of the high-range standard curve is clearly not linear, and our samples, we suspect, are in the below 10 ng/ul point on the curve, the lowest point in the high-range standard curve. So we moved to the low-range curve, as described by Invitrogen, which, in every case failed to yield a good standard curve. We don’t know if it is the assay or perhaps the instrument itself. The low-range standard curve is comprised of the following points for reference [0, 25, and 250 PG/ML, and 2.5 and 25 NG/ML]. We are using a Molecular Dynamics SpectraMax M2 instrument.
So my question is has anyone had success with assaying their ChIP DNA, pre-sample prep., using the PicoGreen assay’s low-range standard curve, particularly with a SpectraMax instrument?
Lastly, has anyone has success performing ChiP-Seq with starting samples in the 4-6 NG range? In our specific experimentation acquiring the 10+ NG of total sample recommended by ILMN has proven difficult.
Any insights would be very much appreciated.
All the best,
Chandi
We’ve been having problems using the PicoGreen assay to determine our final ChIP DNA samples prior to sample prep for use with the ILMN system. While the high-range standard curve is reasonably linear, the lower portion of the high-range standard curve is clearly not linear, and our samples, we suspect, are in the below 10 ng/ul point on the curve, the lowest point in the high-range standard curve. So we moved to the low-range curve, as described by Invitrogen, which, in every case failed to yield a good standard curve. We don’t know if it is the assay or perhaps the instrument itself. The low-range standard curve is comprised of the following points for reference [0, 25, and 250 PG/ML, and 2.5 and 25 NG/ML]. We are using a Molecular Dynamics SpectraMax M2 instrument.
So my question is has anyone had success with assaying their ChIP DNA, pre-sample prep., using the PicoGreen assay’s low-range standard curve, particularly with a SpectraMax instrument?
Lastly, has anyone has success performing ChiP-Seq with starting samples in the 4-6 NG range? In our specific experimentation acquiring the 10+ NG of total sample recommended by ILMN has proven difficult.
Any insights would be very much appreciated.
All the best,
Chandi