Forum: Sample Prep / Library Generation
10-18-2019, 03:42 PM
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Replies: 6
Views: 2,553
The Tween modification plus 6kV for 15sec...
The Tween modification plus 6kV for 15sec injection on the RNA kit has yielded perfect data for the last 4 injections. I am once again satisfied with this instrument, though Agilent tech support was...
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Forum: Sample Prep / Library Generation
10-18-2019, 11:25 AM
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Replies: 6
Views: 2,553
Ran the RNA kit (DNF-471) with an injection of...
Ran the RNA kit (DNF-471) with an injection of 5.5kV for 10sec, and everything worked well. These were samples that had been run previously, and I only loaded 1uL per sample while also bringing the...
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Forum: Sample Prep / Library Generation
10-16-2019, 05:12 PM
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Replies: 6
Views: 2,553
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Forum: Sample Prep / Library Generation
10-16-2019, 11:06 AM
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Replies: 6
Views: 2,553
@weigrc doesn't seem to matter what we run, we...
@weigrc doesn't seem to matter what we run, we just have inconsistent results. The latest saga involves both RAD libraries that were Pippin selected to 200-550nt and raw RNA extracts from cell...
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Forum: Sample Prep / Library Generation
10-15-2019, 01:02 PM
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Replies: 6
Views: 2,553
Fragment Analyzer reliability issues
We acquired our Fragment Analyzer from AATI in March 2018. We primarily use it for NGS library QC (sizing) and to collect RIN values for RNA extracts. At first it was splendid, but within a few...
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Forum: Illumina/Solexa
11-02-2016, 10:55 AM
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Replies: 44
Views: 21,294
Once you have sequenced through adapters, the...
Once you have sequenced through adapters, the garbage sequence that is produced afterward has very low q-scores. If you have adapter/primer sequences that are ~60mer and you are sequencing on a 2x250...
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Forum: Illumina/Solexa
10-27-2016, 11:03 AM
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Replies: 44
Views: 21,294
I don't know of a pub describing this...
I don't know of a pub describing this jackstraw/birds nesting effect, but there was a nice pub last month showing the effect of high cycling conditions on chimera formation...
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Forum: Illumina/Solexa
10-24-2016, 05:02 PM
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Replies: 44
Views: 21,294
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Forum: Illumina/Solexa
10-22-2016, 11:52 AM
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Replies: 44
Views: 21,294
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Forum: Illumina/Solexa
10-22-2016, 11:09 AM
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Replies: 44
Views: 21,294
1100 may be high. Just watch your...
1100 may be high. Just watch your phasing/prephasing. With long inserts you could see loss of data after turn-around.
I ran some ITS inserts recently that had full construct lengths ~650bp with...
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Forum: Illumina/Solexa
08-22-2016, 10:14 AM
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Replies: 44
Views: 21,294
As your insert size increases, you must also...
As your insert size increases, you must also decrease load concentration. For fungal ITS2 or v4-v5 (515-926) inserts I load at 8pM total with the same 30% PhiX.
My understanding is that 12.5pM is...
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Forum: Illumina/Solexa
08-19-2016, 10:51 AM
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Replies: 44
Views: 21,294
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Forum: Bioinformatics
07-06-2016, 10:43 AM
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Replies: 8
Views: 4,423
fastq-join works OK, you just need to relax it...
fastq-join works OK, you just need to relax it because it is so stringent. I use -m 30 -p 30 with good success. Some data is crappier than other data and may need additional grooming before it can be...
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Forum: Illumina/Solexa
12-15-2014, 03:25 PM
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Replies: 55
Views: 32,808
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Forum: Illumina/Solexa
12-15-2014, 03:16 PM
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Replies: 55
Views: 32,808
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Forum: Bioinformatics
06-03-2014, 03:11 PM
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Replies: 23
Views: 6,135
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Forum: Illumina/Solexa
04-04-2014, 05:12 PM
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Replies: 44
Views: 21,294
You are right about the genome-size concern in...
You are right about the genome-size concern in terms of transposase efficiency, but in terms of fragment size, the main factor will be how intact your DNA truly is.
I've never heard the term bird...
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Forum: Illumina/Solexa
04-03-2014, 05:50 PM
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Replies: 44
Views: 21,294
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Forum: Illumina/Solexa
04-03-2014, 05:47 PM
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Replies: 44
Views: 21,294
First, I think I said something slightly...
First, I think I said something slightly incorrectly. I didn't mean to say genome-size dependent, I meant "intact" DNA size dependent. Decent quality DNA extractions yield DNA sheared to around...
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Forum: Illumina/Solexa
04-03-2014, 09:33 AM
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Replies: 44
Views: 21,294
1) I have. It works. Also dependent on genome...
1) I have. It works. Also dependent on genome size.
2) Illumina's recommendation is exactly a 0.5X cleanup. You do incur loss of target DNA at this concentration. Reducing to 0.4X, you could...
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Forum: Illumina/Solexa
04-02-2014, 06:45 PM
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Replies: 44
Views: 21,294
M4TTN: Results looked splendid. Output as...
M4TTN: Results looked splendid. Output as anticipated (17G), ~80%>q30.
I hadn't seen mcnelson's post despite having posted in this thread myself previously, but we also have abandoned the bead...
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Forum: Illumina/Solexa
03-28-2014, 10:53 AM
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Replies: 44
Views: 21,294
austinso, Yaximiks recipe does diulte 100fold...
austinso, Yaximiks recipe does diulte 100fold with hyb buffer. Since he(?) does 3ul NaOH and 3ul of pool then the final dilution achieves this.
Thanks rnaeye for the clarification. I almost...
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Forum: Illumina/Solexa
03-27-2014, 02:20 PM
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Replies: 44
Views: 21,294
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Forum: Illumina/Solexa
03-27-2014, 01:30 PM
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Replies: 44
Views: 21,294
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Forum: Bioinformatics
01-20-2014, 09:18 AM
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Replies: 4
Views: 2,116
I thought I had tried that previously, but the...
I thought I had tried that previously, but the user went and tried the free version of Geneious and used that instead (pretty great assembler). Their lab wound up purchasing a license as did we. If...
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