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Old 04-05-2012, 11:25 AM   #1
beetle_doc
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Default Pippin Prep for TruSeq small RNA libraries

Is anyone using the Pippin Prep to size select their TruSeq small RNA libraries? If so, could you give me information on ease of use, failure rate, and % primer dimer sequences per sample. We are interested in purchasing this instrument and have heard good things about other library types. However, I think that small RNA would be the library that I would run most on this instrument. Any thoughts would be appreciated.
Mike
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Old 04-10-2012, 06:50 AM   #2
mehdik
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Default Pippin Prep conditions for miR-seq libraries

We prepare libraries according to the Illumina small RNA protocol, run them on the bioanalyzer after PCR, quantify the miR band and pool equimass on that band only. Then we use a 3% cassette and set the Pippin to 125-160 collection. You can see from the attached bioanalyzer that the dimers are completely gone. Red is pool pre Pippin, blue is pool post Pippin. It gives great results in my opinion.

The annoying part is that the yield of each individual miR library is different and that it is best to pool after running each library individually on the bioanalyzer. Since V3 at about 200M reads per lane, if you 48plex you get ~4M reads per sample which is a great miR experiment. You might do away with individually quantifying but expect large variations in the number of reads per samples.
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Old 04-16-2012, 07:53 AM   #3
beetle_doc
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Default

That sounds great. What version of Pippin prep are you using?
Mike
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