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12-20-2017, 09:33 AM
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#1 |
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Member
Location: Seoul Korea Join Date: Jan 2014
Posts: 11
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Miseq dual index Read1 (i7 index) problem?
During miseq run, good quaility of read1 and read3 (i5), read4.
But read2 i7 index is low quality. I already found many posting about read3 (i5 index) problem. but I can’t find about i7 index failure. I use Truseq HT dual index primer for 5 sample. Help me please. Last edited by Min Jung Lee; 12-20-2017 at 09:36 AM. |
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12-20-2017, 06:20 PM
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#2 |
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Jafar Jabbari
Location: Melbourne Join Date: Jan 2013
Posts: 1,238
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It could be that i7 indexes are not diverse enough. You can check Illumina pooling guidelines for duel indexed adapters (look at page 21).
http://sapac.support.illumina.com/co...5042173-01.pdf |
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12-20-2017, 06:24 PM
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#3 |
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Member
Location: Seoul Korea Join Date: Jan 2014
Posts: 11
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Thank you for your reply.
but, my library is enough diversity. |
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12-20-2017, 08:28 PM
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#4 |
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Jafar Jabbari
Location: Melbourne Join Date: Jan 2013
Posts: 1,238
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I am pointing to index diversity not the library. I wonder which wells from adapter plate you have used.
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01-08-2018, 12:22 PM
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#5 |
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Senior Member
Location: Purdue University, West Lafayette, Indiana Join Date: Aug 2008
Posts: 2,317
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What sort of adapters are these that you are using?
Illumina instruments that prime i5 using the flow cell oligos (including the MiSeq) will always produce i5 sequence from a library molecule. But to produce an i7 read you need one of the i7 indexing primers to anneal in your amplicon. MiSeq's seem to have a fairly high annealing temp for their sequencing primers... -- Phillip |
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