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Old 02-13-2013, 07:35 AM   #1
casey6r0wn
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Default Nextera-Truseq hybrid library prep

First, many thanks to ECO et al for this thread.

I have a related question.

Is it possible to sequence a library that has a one Truseq end and one Nextera end?

For example:

5'-CommonEnd-Index1-InnerNextera-N^n-InnerTruseq-Index2-CommonEnd-3'

Thanks for you thoughts.
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Old 02-14-2013, 05:13 AM   #2
pmiguel
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Quote:
Originally Posted by casey6r0wn View Post
First, many thanks to ECO et al for this thread.

I have a related question.

Is it possible to sequence a library that has a one Truseq end and one Nextera end?

For example:

5'-CommonEnd-Index1-InnerNextera-N^n-InnerTruseq-Index2-CommonEnd-3'

Thanks for you thoughts.
Presuming you are running the samples on a MiSeq, I think it could work. Devil is in the details, of course...

--
Phillip
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Old 02-14-2013, 05:20 AM   #3
casey6r0wn
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Are you implying that it would not work on a HiSeq, but will on a MiSeq? How will it matter in this context? Thanks for the clarification.

Assuming a library has the correct terminal oligos (for annealing to the flow cell) and one uses the right mix of sequencing primers...
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Old 02-14-2013, 07:30 AM   #4
pmiguel
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Quote:
Originally Posted by casey6r0wn View Post
Are you implying that it would not work on a HiSeq, but will on a MiSeq? How will it matter in this context? Thanks for the clarification.

Assuming a library has the correct terminal oligos (for annealing to the flow cell) and one uses the right mix of sequencing primers...
The MiSeq comes loaded with all the primers needed to sequence both Nextera and TruSeq libraries as well as dual indexes.

The HiSeq requires extra reagents, both primers and SBS reagents to deal with the extra index cycles. At least in "high output mode".

In Rapid mode, I think the situation is similar to that for the MiSeq.

I forgot to note that you still have to set up the sample sheet right for the MiSeq. To get dual indexes to work you have to choose "Amplicon" as your recipe, you get an error if you try to choose "Default". Also, you need to input the index sequences for your i7 and i5 reads.

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