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Forum: Sample Prep / Library Generation 11-22-2018, 12:06 PM
Replies: 17
Views: 9,635
Posted By Seqasaurus
Did you ever get to the bottom of this mchekula?...

Did you ever get to the bottom of this mchekula? We have the exact same from some insect samples
Forum: Sample Prep / Library Generation 03-17-2015, 06:13 AM
Replies: 1
Views: 1,640
Posted By Seqasaurus
sharp peak in library - TruSeq Stranded Total RNA (rRNA-depleted E. coli)

Anyone seen anything like the sharp peaks at 250 bp in the bioanalyzer traces of the libraries below? Overamplified? Bubble product?

These are from rRNA depleted (Ribo zero) total RNA from E....
Forum: Sample Prep / Library Generation 03-12-2014, 02:40 AM
Replies: 4
Views: 3,178
Posted By Seqasaurus
we used these just-a-plate ones a few years back....

we used these just-a-plate ones a few years back. They seem to be the same idea and worked fine as a I remember. http://www.charmbiotech.com/product.html
Forum: Sample Prep / Library Generation 05-13-2013, 05:35 AM
Replies: 1
Views: 2,070
Posted By Seqasaurus
dynabeads/AMPure/SPRIplate compatibility

does anyone know if dynabeads are compatible with SPRIplate magnets for ampure beads or vice versa?
Forum: RNA Sequencing 02-06-2012, 05:33 AM
Replies: 6
Views: 2,044
Posted By Seqasaurus
35 million is my magic number

35 million is my magic number
Forum: Bioinformatics 01-12-2012, 01:07 AM
Replies: 3
Views: 2,039
Posted By Seqasaurus
ml 40 is fine. 1 week isn't long considering you...

ml 40 is fine. 1 week isn't long considering you are using 85 million illumina reads! In the past I used 1.2 million gs flx reads and 2 millions 100bp se hiseq2000 reads for a eukaryote...
Forum: Bioinformatics 11-01-2011, 09:45 AM
Replies: 9
Views: 3,313
Posted By Seqasaurus
For what its worth, i can vouch for newbler 2.6...

For what its worth, i can vouch for newbler 2.6 with 1.2 million titantium plus up to 2million HiSeq 100bp SE. did a good job with eukaryotic cDNA
Forum: Bioinformatics 10-19-2011, 12:06 PM
Replies: 2
Views: 2,634
Posted By Seqasaurus
illumina/454 de novo hybrid cDNA assembly with newbler2.6

Anybody experimented with hybrid assemblies using newbler 2.6?

I fed newbler 1.2 million titanium reads and 2.4 million 100-bp HiSeq2000 v3 reads (a random subset from 12 different treatments of...
Forum: Illumina/Solexa 10-18-2011, 02:24 PM
Replies: 4
Views: 3,638
Posted By Seqasaurus
thanks for the advice. Don't worry, I wasn't...

thanks for the advice. Don't worry, I wasn't planning on wasting an entire lane on one 3-mb genome. There'll be a few bacterial genomes and an insect transcriptome on there too.

that's 4000...
Forum: Illumina/Solexa 10-18-2011, 07:02 AM
Replies: 4
Views: 3,638
Posted By Seqasaurus
paired-end read length for de novo assembly

I've just been told by a sequencing company rep that 50-bp PE reads are "better" for de novo assembly of bacterial genomes than 100-bp PE reads. This seemed dubious to me. At this company a 50-bp PE...
Forum: Bioinformatics 09-20-2011, 11:27 PM
Replies: 2
Views: 1,930
Posted By Seqasaurus
I guess one option would be to assemble illumina...

I guess one option would be to assemble illumina and 454 together in newbler with the -rip option and then to parse the ACE file. However I have ~90 million 100-bp HiSeq reads and so a hybrid...
Forum: Bioinformatics 09-18-2011, 01:19 PM
Replies: 2
Views: 1,930
Posted By Seqasaurus
making non-redundant isotig file from newbler output

Does anyone have decent strategy for taking isotigs.fna (cDNA sequences) and outputting a list of non-redundant isotigs that could be used for mapping rnaseq reads for differential expression...
Forum: Sample Prep / Library Generation 05-31-2011, 12:49 PM
Replies: 8
Views: 4,562
Posted By Seqasaurus
Uppsala Genome Centre plans to start sometime in...

Uppsala Genome Centre plans to start sometime in the autumn
Forum: Literature Watch 05-29-2011, 01:59 AM
Replies: 17
Views: 8,634
Posted By Seqasaurus
regarding the actual subject of the thread, does...

regarding the actual subject of the thread, does anyone know where to get $-750 ion torrent sequencing? (see table 2)
Forum: Ion Torrent 05-29-2011, 01:56 AM
Replies: 14
Views: 9,120
Posted By Seqasaurus
in "Field guide to next-generation DNAsequencers"...

in "Field guide to next-generation DNAsequencers" Glenn lists the minimum cost of an ion torrent run (a full 314 chip) at $750 ("Typical full cost (i.e. including labour, service contract, etc.) of...
Forum: Bioinformatics 05-24-2011, 08:04 AM
Replies: 4
Views: 2,073
Posted By Seqasaurus
transcriptome -> predicted peptide database

can anyone recommend a pipeline to process transcriptome data into a predicted tryptic peptide database?

i.e. we want to do some LC-MS/MS and iTRAQ-MS/MS with our organism however the only...
Forum: General 05-17-2011, 02:00 AM
Replies: 37
Views: 18,592
Posted By Seqasaurus
they've had the 1K upgrade at the CGR liverpool...

they've had the 1K upgrade at the CGR liverpool for almost a year now. Last I heard the average read length was still under 600 in their hands but this may have improved recently
Forum: Bioinformatics 03-28-2011, 02:38 PM
Replies: 6
Views: 2,981
Posted By Seqasaurus
pyfasta http://pypi.python.org/pypi/pyfasta/ ...

pyfasta http://pypi.python.org/pypi/pyfasta/

or flatten the fasta so that each sequence only occupies a single line then use grep?
Forum: RNA Sequencing 03-05-2011, 10:07 AM
Replies: 23
Views: 18,605
Posted By Seqasaurus
I'd have thought polyA selection would have been...

I'd have thought polyA selection would have been the main source of 3' bias. Maybe also fragmentation of cDNA. On 454 at least, cDNAs of a certain size range don't nebulize well. This would bias...
Forum: 454 Pyrosequencing 12-09-2010, 03:04 AM
Replies: 3
Views: 2,788
Posted By Seqasaurus
poor amplification with fusion primers

Hi

we're having trouble with some of our barcoded 16s primers. We are using 27f(fused with primer B) and 519r (fused with MID and primer A). These exact primers have been used before by others (Wu...
Forum: 454 Pyrosequencing 11-02-2010, 03:37 AM
Replies: 1
Views: 1,434
Posted By Seqasaurus
just in case anyone else is interested, I found...

just in case anyone else is interested, I found an alternative a similar kit here http://www.charmbiotech.com/PCR-Norm.htm
Forum: 454 Pyrosequencing 10-26-2010, 05:30 AM
Replies: 1
Views: 1,434
Posted By Seqasaurus
SequelPrep alternative

Can anyone think of an alternative to SequelPrep normalization? i.e. a kit that allows purification and normalization in a single step. There's a delay and SequelPrep isn't available in the UK until...
Forum: Sample Prep / Library Generation 09-07-2010, 10:30 AM
Replies: 4
Views: 3,178
Posted By Seqasaurus
sequalprep normalization of amplicons for titanium

Hi

Has anyone used an invitrogen sequalprep normalization plate to normalize amplicons for multiplexing on 454 titanium? I noticed it was used by Kirk-Harris et al (Applied and Environmental...
Forum: 454 Pyrosequencing 09-07-2010, 03:17 AM
Replies: 2
Views: 1,663
Posted By Seqasaurus
calculating expected reads

Hi,

we are currently trying to decide how to process our 16S amplicons on titanium. How would I come to a rough estimate of expected reads per sample on 1/8th PTP at 16-plex? (assuming everything...
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