A few questions about DNA and isolation/viscosity!

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seabreeze
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A few questions about DNA and isolation/viscosity!

Post by seabreeze » Wed Apr 09, 2008 7:26 am

I am trying to complete an assignment about DNA, and the following questions are nowhere to be found in my textbook/notes nor in any easy-to-understand journals! so any help would be greatly appreciated :smile:

- The difference between DNA and RNA that allows them to be separated...
I know that DNA has a deoxy portion on the ribose, and is double stranded instead of single, but is this what allows it to be soluble and separable in certain solvents that RNA cant? and if so how exactly?
- what base does to DNA and how it affects its viscosity if at all...does it base hydrolyze it or? this one's really confusing me
- what acid does to DNA and how it affects its viscosity if at all...again same as above...really confused
- DNAse affect on visocity of DNA ... this one I know that DNase cleaves and cuts the DNA into parts, and that it reduces its viscosity...but how? it is simply because the molecule is in many smaller parts so it can flow faster or something like that?
- how is DNA precipitated....is it like denatured with certain substances? or is there some sort of charge issue?

Of course I dont expect input on all of these, but any would be really helpful. Thanks!!!

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mith
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Post by mith » Wed Apr 09, 2008 2:53 pm

Think tertiary structure and charges.

hint: the salts act as ligands.

P.S. at microfluidic scale, DNA is pretty large and tends to tangle
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seabreeze
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Re: A few questions about DNA and isolation/viscosity!

Post by seabreeze » Wed Apr 09, 2008 10:30 pm

ok i figured just about all of it out but
can someone please explain which difference between DNA and RNA allows for their separation?

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Re: A few questions about DNA and isolation/viscosity!

Post by blcr11 » Thu Apr 10, 2008 4:42 pm

This pertains mostly to separations by phenol extraction. The ribose ring of RNA is susceptible to elimination/hydroysis reactions at high pH. The deoxyribose ring of DNA is much more stable toward base compared to RNA. At pHs above 7, both RNA and DNA partition into the aqueos phase of a phenol extration, so, when you’re going after DNA and aim to get rid of the RNA, you’ve got to treat the sample with RNase before extraction at pH 8, typically. Then the proteins will denature and float at the interface, the small mw ribonucleotides will preferentially partition into the organic phase, leaving you with DNA (which is stable toward high pH) in the aqueous phase. RNA is much more stable toward low pH than high. At low pH, DNA is denatured. So doing the extraction at pH 4.5 will leave only RNA in the aqueous phase, the DNA and protein all partitioning to the organic phase or precipate at the interface.

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