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Old 08-22-2013, 08:02 AM   #1
miRman
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Default Ion Torrent RNA library prep

Can anybody here explain the molecular biology behind Ion Torrent RNA library prep? The protocol is particularly vague about adapter hybridization and ligation which is what I would like to understand.

According to the protocol, first the adapters are annealed and then ligated. Does this mean that the adapters are partly double stranded or a hairpin?
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Old 08-28-2013, 03:55 AM   #2
miRman
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I found out some details of the prep from Ion Torrent directly. It seems that there are two distinct oligos per fragment end.

See http://www.labome.com/method/RNA-seq...equencing.html for a flow diagram describing the library prep and more details.
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Old 04-30-2014, 11:25 AM   #3
mbzmg1
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Correct -the adapters that are hybridized and ligated to your RNA fragments have degenerate overhangs that allow for strand-specific ligation, and therefore sequencing. As far as I can gather this is essentially the same strategy as older Ambion RNAseq kits (eg http://www.lifetechnologies.com/us/e...equencing.html).

See also the attached slide.

Hope this helps.
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