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Old 01-07-2013, 04:19 AM   #1
SLeigh-Brown
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Location: Cambridge, UK

Join Date: Dec 2011
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Question Low cluster density of Truseq "high throughput" libraries - both DNA and RNA

Hi genomics folks,

Has anyone got any experience in clustering Illumina Truseq RNA 'HT' (high throughput) and Truseq DNA HT libraries?

I work in an NGS service, and we recently tried out the new Truseq HT kits. I like the idea of multiplexing up to 96 Truseq libraries within a single lane.

These libraries were quantified by qPCR following our normal protocol for Truseq libraries, using the KAPA library quant kit. They were then clustered using a cbot -> HiSeq or straight on to the MiSeq. The resulting cluster density was 1/4 to 1/2 of what we would expect, on both machines.

These projects were unrelated and the library prep was performed by very experienced guys in the lab - I'm starting to wonder if the HT kits require different clustering conditions. According to Illumina, these libraries should cluster OK using the normal protocol.

Has anyone else experienced this problem?
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cluster density, library quantification, truseq high throughput

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