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Labelling ChIP samples - ARG! Hope someone can help! (May/06/2008 )

Hi smile.gif

And so we thought we were getting close to getting our chip-chip sorted...
It turns out our hybridisation issues may be because of our labelled samples. ie: when we look at the green and red intensities (using Agilents Feature Extraction), they are quite low (about 4000). Another group here has been getting about 40, 000! So I wonder if our labelling reactions are crap? We have tried two methods: using Invitrogen's "BioPrime total genomic kit" and the "BioPrime Kit" that use exo-klenow and klenow respectively. With Klenow we end up with more DNA but the amount of dye incorporation is similiar between the two kits (~150pmol total dye incorporation for each sample).
Can anyone give us some advice?
Thanks in advance!


I *think* I have solved the problem. Instead of eluting our amplified samples in Tris - I eluted in water and YAY! - the degree of labeling increased biggrin.gif
Now onto the next hybridisation.....

Have a great weekend everyone!