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Old 01-28-2013, 05:52 AM   #1
addyblanch
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Default Illumina Nextera Enrichment Buffers

Hi Everyone,

Just a quick question.

I'm currently trying to enrich a library prep using solution hybrid capture. I'm following the Nextera Enrichment protocol however because i'm studying bacteria I cannot use the kit itself.

I'm just wondering if anyone knows what the buffers ET1 and HP3 contain? They are used to elute samples from a streptavidin/biotin complex. I'm guessing they denature the DNA so its released from the biotinylated primer.

If not does anyone have a suggestion that would be fit to use before sequencing on a MiSeq.

Adam
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Old 02-04-2014, 10:45 AM   #2
cement_head
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Quote:
Originally Posted by addyblanch View Post
Hi Everyone,

Just a quick question.

I'm currently trying to enrich a library prep using solution hybrid capture. I'm following the Nextera Enrichment protocol however because i'm studying bacteria I cannot use the kit itself.

I'm just wondering if anyone knows what the buffers ET1 and HP3 contain? They are used to elute samples from a streptavidin/biotin complex. I'm guessing they denature the DNA so its released from the biotinylated primer.

If not does anyone have a suggestion that would be fit to use before sequencing on a MiSeq.

Adam
HP3 = 2.0 N NaOH
ET1 = ?

Last edited by cement_head; 12-16-2014 at 08:10 AM.
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Old 02-04-2014, 12:53 PM   #3
buckybadger
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The selection of which protocol to use will be dependent on the library prep and the type of probe, as well as (but to a lesser degree) the size of the target, the sample type (e.g. pure bacterial genomic DNA or DNA from bacteria mixed with a complex metagenomic or host sample?), and how you intend to sequence and analyze that sequence.

Was your library made using the Nextera kit, or do you want to enrich a non-Nextera library but just using the Nextera capture protocol? If it is a Nextera library, was it made using the Nextera DNA Sample Preparation Kit or one of the Nextera Enrichment kits?

Are you planning to enrich one sample at a time or planning a multiplex capture? The latter requires indexed libraries if you need to preserve sample identification information, while the former may or may not require indexing depending on whether or not you want to multiplex for sequencing but not for capture.

What kind of biotinylated probes are you planning to use...RNA or DNA?
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