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Does small amount of RNA affect MeDIP? - (Jul/20/2008 )

QUOTE (cjbhm @ Jul 20 2008, 07:26 AM)
Hi, I am doing MeDIP. After purification with Qiagen's Allprep DNA/RNA mini kit (just follow the typical procedure without treating with RNase A), there may be a very small amount of RNA in my DNA samples (OD 260/280=1.84 to 1.87). But it seems not affect the sonication as showed in the attached gel pics. So do I need to treat my DNA samples after or before sonication? mellow.gif I can not find any evidence that small amount RNA will affect IP. After IP, there should be no RNA to affect microarray.

Pic 1, 2% gel, 67V 20min. Pic 2, same gel runing another 20min. Pic 3, stay in TBE runing buffer for 2 hr. Sonicated on Sonics VCX 130, 2ug DNA in 200ul TE. Load 18ul. The marker is NEB's 100bp ladder: 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1200, 1517
1, 30% power, 10s on, 40s off, 10 times
2, 30% power, 20s on, 40s off, 10 times
3, 30% power, 30s on, 40s off, 10 times
4, 40% power, 20s on, 40s off, 10 times
5, 6, 20% power 5s on10s off. 5-10 times

Which one can be used on IP? 2 is 150-800bp and 3 is 150-600bp?
What is the 200bp around belt in Pic 2? RNA? There is no belt in pic 1, and they disappeared after 2 hrs later. I also did not find on DNA samples without sonication.

-cjbhm-

I find RNA do affect MeDIP from http://www.epigenome-noe.net/researchtools...l.php?protid=33 And they use big fragment similar to line 1. However I am trying conditions between line 2 and line 3 and hope to make peak more defined.

-cjbhm-

I find RNA do affect MeDIP from http://www.epigenome-noe.net/researchtools...l.php?protid=33 And they use big fragment similar to line 1. However I am trydefined. ing conditions between line 2 and line 3 and hope to make peak more defined.

-cjbhm-

sorry, always send two times

-cjbhm-