Genomic DNA smears on gel - (Jan/25/2008 )
I'm currently purifying genomic DNA from my Gram negative bacteria using the kit from Promega (Wizard Genomic DNA Purification) but I keep getting smears when visualized. The manufacturer's protocol states the use of 3uL of RNase. Just thinnking: could I increase the amount? Are those smears caused by RNAs?
Things turn out exactly to be like in the manual: especially thread-like mass of DNA, but not getting single bright band.
Loaded 5uL of sample + 1uL of 6X loading dye. Hope you guys could give input. Thanks.
Dont worry about the RNA. They don't cause smearing.
I will suspect either protein contamination, degradation (which is quite unlikely) or too concentrated DNA sample.
My suggestion is to wash it thoroughly to get rid of unwanted protein in it.
Hope this help.
Oh, it will be better if you can show us the gel picture as well.
I will upload it soon. Say I have 100uL of DNA samples now, do I redo or can I clean it? Lets see, spin it down? I'm really drained. Thanks Tim.
Maybe dilute the samples at run them again. A couple of dozen ng may be enough to gel-check the DNA.
I use that kit and it works brilliantly, although I only load 1ul on a gel to analyse this, is suspect more will make it smear. Is the smear from the top pf the gel down or in a discrete part of the gel?
From the top down. What's the most-used amount to load in the gel? Perhaps my 5uL + 1uL dye is too much. Will work on it again.