DNA Gel Extraction Problem - multiple bandings in gel extraction product (Sep/16/2005 )

Hi,

I've recently encountered recurring problems with my DNA gel extraction. I would run a 1% agarose gel, then excise the specific band and do the gel extraction using a kit. But when I check the results I get multiple bandings!
Can someone help me on this? Could it be because of overloading the gel?
Thanks a lot.

Rgds,
Sherene

There are many reasons for the problem. Upload your gel picture.

1. DNA extraction from agarose gel does not guarentee 100% purification of the desired-sized DNA. Overloading causes this problem, too. If you got the same bands to the original ones, and the not-desired bands got weaker, i would say, it happens. Just re-purify the desired band. Or increase or decrease the agarose percentage to adjust the condition for easy separation.

2. Did you check the buffers from the kit, including your water?


.

You may denatured your DNA - this can give multiple bands.

Daniel

DNA sequencing software

>>I had same problem. When I decreased quantity of DNA in my sample, it became better.
>> In some cases decreasing % of agarose downto ~0.8 solve this problem

Hi,
Firs of all, What is the size of your DNA? This is so important, If the size of DNA bigger than 6kb, you must use 0.8-0.7% agarose gel. I think that your DNA degraded. At this point, you must run the DNA slowly and your purification kit is also important. I use the Applichem DNA isolation Kit and you work well.
Good luck...

Hi all,

Thank you so much for all your input. Sorry I forgot to mention that the multiple bandings I got were actually multiply larger than my desired band, and yes, the bands were weaker than the desired band (but still a nuisance nonetheless!). So, I doubt that the problem is degradation of DNA.

Anyway, I tried to use 1.2% agarose, made sure I didn't overload this time and ran the gel longer (40 min at 120V) to achieve better separation and get a tighter band and avoid overlapping fragments, and hey, this time it worked! I'm so happy...

God bless you.

swyin.