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Forum: Illumina/Solexa 01-28-2014, 08:27 AM
Replies: 10
Views: 4,613
Posted By bstamps
Well, once again it depends. Sure, V3/V4...

Well, once again it depends. Sure, V3/V4 sequencing will allow you to ID the microbial community to the level of family (Maybe genus), which given enough samples and experimental variables would...
Forum: Illumina/Solexa 01-28-2014, 05:50 AM
Replies: 10
Views: 4,613
Posted By bstamps
1. How much depth is sample dependent. Do you...

1. How much depth is sample dependent. Do you have a hypothesis as to the diversity of the environment? Do you have a question/hypothesis that can be tested with more/less depth? 3 million reads per...
Forum: Metagenomics 01-20-2014, 02:40 PM
Replies: 8
Views: 13,871
Posted By bstamps
You know there is a pre-existing way to have...

You know there is a pre-existing way to have multiple runs of 454, Illumina...whatever... within QIIME, right? Just add -n some integer to the split_libraries.py or split_libraries_fastq.py that...
Forum: Bioinformatics 01-15-2014, 01:53 PM
Replies: 8
Views: 2,584
Posted By bstamps
Give it a shot- I've never had issues with...

Give it a shot- I've never had issues with App-Deploy before. The last script executed is the activation of the qiime-1.8.0-release/ directory, so if there are any other failures prior, you'll still...
Forum: Bioinformatics 01-15-2014, 05:49 AM
Replies: 8
Views: 2,584
Posted By bstamps
Do you need QIIME 1.7 still functional? Basically...

Do you need QIIME 1.7 still functional? Basically if QIIME was installed by its defaults (~/qiime_software/) it should update any packages inside of this directory, install a qiime-1.8.0-release/...
Forum: Bioinformatics 01-15-2014, 05:32 AM
Replies: 8
Views: 2,584
Posted By bstamps
Have you tried QIIME-Deploy?...

Have you tried QIIME-Deploy? https://github.com/qiime/qiime-deploy

Relatively simple way to upgrade/install QIIME on any Linux box.
Forum: Sample Prep / Library Generation 11-02-2013, 07:06 AM
Replies: 4
Views: 7,874
Posted By bstamps
The instrument also lets you "calculate stock...

The instrument also lets you "calculate stock concentration" for each sample. Just click the "Calculate Stock Concentration" button after reading a sample, set the units to ng/uL (Or whatever else...
Forum: Sample Prep / Library Generation 11-02-2013, 06:20 AM
Replies: 4
Views: 7,874
Posted By bstamps
Well, from the Life Tech website... The BR...

Well, from the Life Tech website...

The BR kit has a range of 100pg/uL to 1000ng/uL and the HS has a range of 10pg/uL to 100ng/uL. If you're on the extreme of the BR kit (Sample is 0.150 ish...
Forum: The Pipeline 10-18-2013, 10:37 AM
Replies: 80
Views: 197,067
Posted By bstamps
I'm guessing A) 100% B) 75/25. Either...

I'm guessing


A) 100%
B) 75/25. Either they release something, or risk losing funding from investors at this point. My bet is that they release the Minion sooner than the Gridion.
Forum: Illumina/Solexa 10-09-2013, 01:03 PM
Replies: 5
Views: 1,995
Posted By bstamps
Could you share how much it was? I'm curious how...

Could you share how much it was? I'm curious how it compares to the LifeTechnologies magnets.
Forum: Sample Prep / Library Generation 10-02-2013, 11:50 AM
Replies: 1
Views: 752
Posted By bstamps
Quantifiable or no? The QuBit will quantify...

Quantifiable or no?

The QuBit will quantify down to 250 pg/uL RNA...but if you just want to know it's there I suppose you could just perform RT/PCR with some marker gene.
Forum: General 09-13-2013, 07:43 AM
Replies: 0
Views: 1,791
Posted By bstamps
Submission of genome annotated via RAST to GenBank

Hello all,

I've been wrestling with this issue the past week... I have a completed (60ish contigs) bacterial genome that I annotated using RAST. I have the .gbk file from this annotation and the...
Forum: Metagenomics 09-06-2013, 07:49 AM
Replies: 1
Views: 2,210
Posted By bstamps
what are you using to do all of this? I'm...

what are you using to do all of this? I'm assuming QIIME by the steps you listed so far.

Check out

http://qiime.org/tutorials/tutorial.html
Forum: Illumina/Solexa 08-20-2013, 06:17 AM
Replies: 26
Views: 14,262
Posted By bstamps
I'd love to see the Qscore plot on that run....

I'd love to see the Qscore plot on that run. Excited for amplicon sequencing to see it move out to 2x300. Anyone know the expected release date for said kit (Though I guess that was the question of...
Forum: Metagenomics 08-14-2013, 06:08 AM
Replies: 9
Views: 5,722
Posted By bstamps
Honestly? Fermentas Taq. Nothing special- when...

Honestly? Fermentas Taq. Nothing special- when you think about it you're generating a 300-500bp fragment, and the likelihood of a misincorporation is low enough that I can live with it. It works well...
Forum: Pacific Biosciences 08-07-2013, 04:08 AM
Replies: 3
Views: 2,700
Posted By bstamps
If you plan on only amplifying 500bp, I might ask...

If you plan on only amplifying 500bp, I might ask why you aren't using Illumina and overlapping each read 50bp and then stitching the reads to get a final artificial read near 500bp (Or waiting a few...
Forum: RNA Sequencing 05-11-2013, 10:16 AM
Replies: 5
Views: 2,198
Posted By bstamps
Trying again, converted my sorted.bam to a...

Trying again, converted my sorted.bam to a sorted.sam and had this error crop up.

Warning: Read D3NH4HQ1:160:C1UJ8ACXX:7:1111:18163:68474 claims to have an aligned mate which could not be found....
Forum: RNA Sequencing 05-11-2013, 07:41 AM
Replies: 5
Views: 2,198
Posted By bstamps
That did the trick. New problem... ...

That did the trick.

New problem...

Warning: No features of type 'exon' found.
Warning: Malformed SAM line: MRNM != '*' although flag bit &0x0008 set
Warning: Malformed SAM line: RNAME != '*'...
Forum: RNA Sequencing 05-11-2013, 07:22 AM
Replies: 5
Views: 2,198
Posted By bstamps
HT-Seq Count (Banging Head Against Wall)

So I've seen this error crop up in the forums several times, but attempting to solve with the suggestions (find/replace within the gff file, use the -i option) doesn't seem to help...

When I run ...
Forum: Metagenomics 03-26-2013, 07:36 PM
Replies: 4
Views: 4,740
Posted By bstamps
1) If this is a single MiSeq run, and the entire...

1) If this is a single MiSeq run, and the entire run was devoted to 16s, expect to use at or greater than 32GB of RAM. Our collaborators attempted to do this on the QIIME virtualbox, and had it crash...
Forum: Vendor Forum 03-05-2013, 06:54 AM
Replies: 5
Views: 4,115
Posted By bstamps
My lab would be interested if you had a kit for...

My lab would be interested if you had a kit for the MiSeq. Sent an email to your customer service- if you need willing participants I'd love to try a few environmental libraries.
Forum: Illumina/Solexa 02-22-2013, 03:44 PM
Replies: 1
Views: 1,400
Posted By bstamps
One I'd ask what primer set you used, and what...

One I'd ask what primer set you used, and what hypervariable regions you cross. You could artificially insert NNNNNNN between the reads (A bad idea in my opinion), though really you might be just as...
Forum: Illumina/Solexa 02-18-2013, 01:37 PM
Replies: 32
Views: 16,740
Posted By bstamps
I'll say that we've had good success having 50%...

I'll say that we've had good success having 50% genomic DNA of an organism we needed sequenced anyway/felt like getting data on, and having our amplicon library with a 12bp random barcode on the...
Forum: Illumina/Solexa 01-25-2013, 08:10 AM
Replies: 11
Views: 3,187
Posted By bstamps
By pre-sheared I think he means your amplicon is...

By pre-sheared I think he means your amplicon is already to an insert size that is acceptable to library prep- all fragments are roughly the same size (It being amplicon), so all you need to do is...
Forum: De novo discovery 01-25-2013, 06:04 AM
Replies: 8
Views: 8,049
Posted By bstamps
We run our analyses on a 334 node Intel cluster...

We run our analyses on a 334 node Intel cluster (2x Xeon [email protected] 2.0 GHz with 32 GB RAM per node)- I usually call 10 nodes at a time, though I work with bacterial genomes...it takes around 15 minutes to...
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