Forum: Sample Prep / Library Generation
04-01-2020, 10:24 AM
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Replies: 4
Views: 2,410
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Forum: Sample Prep / Library Generation
03-31-2020, 04:36 AM
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Replies: 4
Views: 2,410
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Forum: Sample Prep / Library Generation
03-09-2018, 01:47 PM
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Replies: 3
Views: 3,839
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Forum: Illumina/Solexa
01-29-2018, 03:39 PM
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Replies: 2
Views: 1,373
Libraries over 2 kb don't tend to work on...
Libraries over 2 kb don't tend to work on Illumina sequencers, so do you really care what the concentration of those molecules is?
Also, are you doing a TaqMan qPCR or SYBR? If it's SYBR, you have...
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Forum: Illumina/Solexa
01-29-2018, 03:34 PM
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Replies: 1
Views: 1,460
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Forum: Bioinformatics
11-13-2017, 04:17 PM
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Replies: 4
Views: 1,431
If enzymes count as chemicals, NEB has its...
If enzymes count as chemicals, NEB has its "Fragmentase" mix, which appears to be a combination of a dsDNA nicking enzyme that's had its sequence specificity engineered out of it and a second enzyme...
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Forum: RNA Sequencing
11-03-2017, 04:26 PM
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Replies: 4
Views: 4,809
No, I'm concerned that having a lot of random...
No, I'm concerned that having a lot of random bases would invite mispriming and other molecular tomfoolery, plus it's a waste of sequencing cycles if you're using short reads, as we do. Oh, and...
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Forum: RNA Sequencing
11-02-2017, 08:15 AM
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Replies: 4
Views: 4,809
Both great questions.
Well, first of...
Both great questions.
Well, first of all, it has nothing to do with streptavidin, which always confuses people, and that's why our diagram (fig. S1 in Supplemental File 1) just calls it a...
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Forum: RNA Sequencing
11-01-2017, 10:24 AM
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Replies: 4
Views: 4,809
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Forum: Illumina/Solexa
05-24-2017, 10:51 AM
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Replies: 23
Views: 10,123
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Forum: Sample Prep / Library Generation
06-03-2016, 01:01 PM
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Replies: 3
Views: 3,411
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Forum: Sample Prep / Library Generation
03-15-2016, 08:39 AM
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Replies: 7
Views: 3,317
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Forum: Sample Prep / Library Generation
03-08-2016, 06:43 PM
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Replies: 289
Views: 155,147
No, my design was somewhat different. I remain...
No, my design was somewhat different. I remain skeptical of LNAs as discussed previously in this thread. As also discussed previously, Exiqon is the only vendor of LNAs in North America.
But if...
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Forum: Illumina/Solexa
01-24-2016, 12:08 PM
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Replies: 31
Views: 12,051
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Forum: Illumina/Solexa
01-22-2016, 03:03 PM
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Replies: 31
Views: 12,051
Okay, so basically no cost-per-read advantage...
Okay, so basically no cost-per-read advantage over the MiSeq either. I don't mind paying an extra $50 per run to double my read lengths. But if I were buying a new machine, that wouldn't be worth the...
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Forum: Illumina/Solexa
01-21-2016, 12:41 PM
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Replies: 31
Views: 12,051
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Forum: Illumina/Solexa
12-02-2015, 10:14 AM
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Replies: 12
Views: 3,542
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Forum: Sample Prep / Library Generation
12-02-2015, 08:22 AM
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Replies: 23
Views: 8,582
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Forum: Sample Prep / Library Generation
12-02-2015, 07:32 AM
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Replies: 23
Views: 8,582
Is this a typo? Even starting from a single...
Is this a typo? Even starting from a single molecule, 70 PCR cycles would give you about 21 g of DNA, according to a quick back-of-the-envelope calculation, though of course you'll exhaust the...
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Forum: Sample Prep / Library Generation
12-02-2015, 07:14 AM
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Replies: 23
Views: 8,582
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Forum: Illumina/Solexa
12-02-2015, 07:02 AM
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Replies: 12
Views: 3,542
Your .docx protocol says you can follow the...
Your .docx protocol says you can follow the standard MiSeq protocol, so I would do that. Illumina's .pdf protocol has a few special instructions to watch out for. I'm fairly certain the v2 kits...
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Forum: Illumina/Solexa
12-01-2015, 10:47 AM
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Replies: 12
Views: 3,542
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Forum: Illumina/Solexa
12-01-2015, 08:20 AM
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Replies: 12
Views: 3,542
This is just the older version of the standard...
This is just the older version of the standard protocol (for step 8 above). For the current MiSeq v3 kits, you start with 5 uL @ 4 nM, then add NaOH to denature it, then dilute this mix to 1000 uL...
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Forum: Illumina/Solexa
12-01-2015, 07:13 AM
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Replies: 12
Views: 3,542
Oh, so you're asking about a third thing, the...
Oh, so you're asking about a third thing, the pooling after library prep but before target enrichment.
The steps are, roughly:
Isolate 50 ng gDNA from each sample
Normalize each gDNA sample...
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Forum: Illumina/Solexa
12-01-2015, 05:49 AM
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Replies: 12
Views: 3,542
Are you asking about the amount of DNA you use...
Are you asking about the amount of DNA you use for library prep, or the amount of library you use for sequencing?
The final amount of library you use for sequencing is 5 uL @ 4 nM, at least on a...
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