How long the template can be stored at -20?

illuminaGA · 07-08-2013, 07:16 AM

Dear everyone.

I have diluted my libraries to 12pM templates about 2 weeks ago ,and stored at -20 degree. Can I use those templates for cluster generation today? Or I must use fresh diluted templates?

Thank you so much!

TonyBrooks · 07-08-2013, 07:46 AM

We've re-run 12pM a couple of times before after 2 weeks. The data was fine but we did see a marginal reduction in cluster density. We don't do it often enough to say with certainty that it'd be OK though.
We've kept 20pM for 2 months and that seemed to be fine too. In fact we now use a 20pM stock to prepare our PhiX MiSeq spike-ins and just make up some fresh 20pM every few weeks.

memento · 07-17-2013, 10:52 AM

Based on TruSeq library, You absolutely have to run it the same day after multiplexing is finished. This will ensure ~800k/mm2 clusters. We noticed that running the very same sample (stored on SGP, non-diluted) after just 4 days results in a drop from 850k to ~500k. After two weeks we had results ~350k/mm2...Im not sure if there is anything that we can do - overloading works up to a limit of NaOH concentration.

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07-08-2013, 07:16 AM	#1
illuminaGA Member Location: Atlanta Join Date: Dec 2012 Posts: 70	How long the template can be stored at -20? Dear everyone. I have diluted my libraries to 12pM templates about 2 weeks ago ,and stored at -20 degree. Can I use those templates for cluster generation today? Or I must use fresh diluted templates? Thank you so much!

07-08-2013, 07:46 AM	#2
TonyBrooks Senior Member Location: London Join Date: Jun 2009 Posts: 298	We've re-run 12pM a couple of times before after 2 weeks. The data was fine but we did see a marginal reduction in cluster density. We don't do it often enough to say with certainty that it'd be OK though. We've kept 20pM for 2 months and that seemed to be fine too. In fact we now use a 20pM stock to prepare our PhiX MiSeq spike-ins and just make up some fresh 20pM every few weeks.

07-17-2013, 10:52 AM	#3
memento Member Location: UK Join Date: Nov 2010 Posts: 49	Based on TruSeq library, You absolutely have to run it the same day after multiplexing is finished. This will ensure ~800k/mm2 clusters. We noticed that running the very same sample (stored on SGP, non-diluted) after just 4 days results in a drop from 850k to ~500k. After two weeks we had results ~350k/mm2...Im not sure if there is anything that we can do - overloading works up to a limit of NaOH concentration.