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Old 05-23-2012, 03:09 AM   #1
CHE
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Location: London UK

Join Date: May 2012
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Default RNA fragmentation VS cDNA size selection

Hello,

I am relatively new to RNA-seq library synthesis so forgive a potentially naive question.

I have seen a lot of posts discussing fragemtnation of RNA as a method of size selection prior to cDNA synthesis. However this seems like a lot of extra steps at the RNA stage rather than a conversion of RNA to cDNA then fragmentation with size selection. Can anyone tell me the wisdom behind this?

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Chris
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Old 05-24-2012, 11:53 AM   #2
pmiguel
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Location: Purdue University, West Lafayette, Indiana

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RNA can be fragmented using moderate heat/divalent cation solutions. Hence you can quickly fragment 96 samples in a thermal cycler. Fragmenting DNA using non-enzymatic methods is much more laborious.

Enzymes tend to have much stronger sequence-specific biases than chemical/mechanical fragmentation methods, and are less favored for that reason.

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Phillip
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