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Forum: Illumina/Solexa 06-13-2017, 02:38 PM
Replies: 2
Views: 1,057
Posted By kmkocot
Thanks for the quick reply. For what it's worth,...

Thanks for the quick reply. For what it's worth, I measured concentration by Qubit and was pretty careful during all steps, especially the clean-up, so I was surprised to see this much variation.
Forum: Illumina/Solexa 06-13-2017, 01:30 PM
Replies: 2
Views: 1,057
Posted By kmkocot
Nextera XT from ds cDNA

Hi all,

I made cDNA libaries using the Clontech SMART-Seq v4 ultra low RNA kit and made Nextera XT sequencing libraries from that cDNA. The bioanalyzer result (DNA HS) for the Nextera XT libraries...
Forum: Illumina/Solexa 11-21-2016, 02:36 PM
Replies: 1
Views: 2,411
Posted By kmkocot
HiSeq 2500 rapid run versus high output

Hi all,

I'm taking advantage of an end-of-the-year RNA-Seq promotion from my sequencing provider. I was given the option of using their U.S. or overseas sequencing center. They said "if we were to...
Forum: Bioinformatics 06-06-2016, 10:30 AM
Replies: 1
Views: 1,138
Posted By kmkocot
ALLPATHS-LG libary prep question

Hi all,

I'm interested in using ALLPATHS-LG for a genome project. I've read their manual (ftp://ftp.broadinstitute.org/pub/crd/ALLPATHS/Release-LG/AllPaths-LG_Manual.pdf) and I'm a bit confused...
Forum: Illumina/Solexa 06-06-2016, 09:09 AM
Replies: 3
Views: 1,469
Posted By kmkocot
Question Do I need 2+ mate pair libraries with different insert sizes?

Hi all,

I would like to sequence and de novo assemble a draft genome from a marine invertebrate with a roughly human-sized genome: 3.46 gigabases (as estimated by FIAD). I know nothing about...
Forum: Bioinformatics 02-20-2016, 01:27 PM
Replies: 4
Views: 1,839
Posted By kmkocot
For what it's worth, I would strongly recommend...

For what it's worth, I would strongly recommend the newest TransDecoder over ESTScan.

I eventually did figure this out but I have no idea how as it has been years. Sorry I'm not more helpful.
...
Forum: General 09-09-2015, 02:15 PM
Replies: 2
Views: 1,250
Posted By kmkocot
Thanks! This was not on my radar but should be...

Thanks! This was not on my radar but should be very helpful!

Best,
Kevin
Forum: General 09-08-2015, 08:39 PM
Replies: 2
Views: 1,250
Posted By kmkocot
Annotating a contig and bacterial or eukaryotic (contamination screening)?

Hi all,

We are sequencing the genome of a microscopic invertebrate. Our assembly has significant bacterial contamination that we would like to remove prior to gene modelling. Is anyone aware of a...
Forum: General 05-20-2015, 09:17 PM
Replies: 1
Views: 1,649
Posted By kmkocot
WGS on a tiny invertebrate: what library prep and assembly strategy would you use?

Hi all,

We are planning to sequence the genome of a tiny marine invertebrate. DNA extraction from a single individual will likely yield only around 10 ng of DNA and we only want to use one...
Forum: Bioinformatics 06-26-2014, 06:34 PM
Replies: 23
Views: 31,545
Posted By kmkocot
I was having the same issue. Specifying the full...

I was having the same issue. Specifying the full path worked for me too. Thanks!

Kevin
Forum: Illumina/Solexa 05-30-2014, 03:16 AM
Replies: 4
Views: 1,449
Posted By kmkocot
Thanks for the input. I neglected to mention this...

Thanks for the input. I neglected to mention this stuff definitely inhibits PCR (DNA has to be diluted to absurdly low concentrations for PCR to work) so I'm guessing it would cause problems in...
Forum: Illumina/Solexa 05-30-2014, 02:12 AM
Replies: 4
Views: 1,449
Posted By kmkocot
DNA extraction for mate pair lib. from pigmented marine inverts

Hi all,

I need to extract DNA from some frustrating marine invertebrates. They are slimy and have a pigment that must have some chemical properties similar to DNA. When I do an extraction with...
Forum: Bioinformatics 05-04-2014, 10:16 PM
Replies: 0
Views: 1,393
Posted By kmkocot
PFAM annotation - need help with awk scripting

Hi all,

I'm using pfam_scan.pl to annotate some sequences from a gene family. I'd like to take the output of my pfam annotation, which looks like this...


# pfam_scan.pl, run at Mon May 5...
Forum: Bioinformatics 04-10-2014, 04:31 PM
Replies: 4
Views: 1,839
Posted By kmkocot
ESTScan 3 installation

Hi all,

I'm having trouble installing ESTScan 3. Anyone have any idea what my problem is?


kmkocot@Epimenia:~/Desktop/estscan-3.0.3$ make
gcc -O2 -c maskred.c
maskred.c: In function...
Forum: Bioinformatics 04-01-2014, 07:23 AM
Replies: 3
Views: 2,026
Posted By kmkocot
Will this count all of the reads in the input...

Will this count all of the reads in the input .fastq files or just the reads that mapped? I think the latter is the case, right? Is there a quick script / tool you can recommend to do this in batch?
Forum: Bioinformatics 03-31-2014, 04:33 PM
Replies: 3
Views: 2,026
Posted By kmkocot
rsem - find number of reads in original data

Hi all,

I ran rsem on a bunch of Illumina transcriptomes to get FPKM values. Because of the large file size, I deleted the .fastq files containing the raw reads from my local machine. Now I...
Forum: Bioinformatics 03-26-2014, 11:15 PM
Replies: 5
Views: 12,620
Posted By kmkocot
Update

Hi all,

I tried again by running the following script:

fastq_quality_filter -q 20 -p 90 -Q33 -i test.fastq | fastq_quality_trimmer -t 20 -l 15 -Q33 | fastx_trimmer -f 11 -m 15 -o test_1.fastq...
Forum: Bioinformatics 03-25-2014, 05:25 AM
Replies: 5
Views: 12,620
Posted By kmkocot
Thanks guys. I'll try your suggestions (less...

Thanks guys. I'll try your suggestions (less strict trimming, more kmers) and report back. Adapters were already trimmed and verified absent with fastqc. I tried a kmer of 21 on untrimmed data and...
Forum: Bioinformatics 03-24-2014, 03:18 AM
Replies: 5
Views: 12,620
Posted By kmkocot
Question Trying to estimate genome size - no peak in jellyfish kmer abundance plot

Hi all,

I have some 2 X 100 PE Illumina genomic data from a marine invertebrate whose genome size I'd like to estimate. I was trying to follow the workflow presented here...
Forum: Illumina/Solexa 10-22-2013, 11:31 AM
Replies: 21
Views: 16,062
Posted By kmkocot
Of 3 tries with 15, 20, and 40 minutes on 8...

Of 3 tries with 15, 20, and 40 minutes on 8 diverse invertebrates (extracted with the Qiagen DNEasy kit) only one worked. The rest looked like unfragmented gDNA after even 40 minutes.
Forum: General 09-17-2013, 07:00 AM
Replies: 0
Views: 2,411
Posted By kmkocot
Question Pooling and diluting Nextera libraries

Hi all,

I made some Nextera libraries last week and wanted to pool 8 of them for sequencing. Here's the Qubit concentration and volume containing 2 nM for each given the equation 1ng/ul = 3nM...
Forum: Bioinformatics 05-14-2013, 05:53 PM
Replies: 2
Views: 2,390
Posted By kmkocot
Thanks! I'm testing it now.

Thanks! I'm testing it now.
Forum: Bioinformatics 05-14-2013, 03:41 PM
Replies: 2
Views: 2,390
Posted By kmkocot
Easiest way to get number of reads per contig?

Hi all,

I'm trying to map 2 X 100 PE Illumina reads (transcriptome) back to a small number of contigs. I want to get the number of reads that align to each contig as quickly as possible. Here's...
Forum: Bioinformatics 02-25-2013, 04:24 PM
Replies: 7
Views: 6,109
Posted By kmkocot
Unfortunately I didn't figure out my issue. I...

Unfortunately I didn't figure out my issue. I will probably work with this some more in the not too distant future so if I figure anything out I will let you know.

Kevin
Forum: Bioinformatics 02-25-2013, 02:55 PM
Replies: 278
Views: 117,431
Posted By kmkocot
Me again. seb567, the link to the visualization...

Me again. seb567, the link to the visualization tool you posted (http://genome.ulaval.ca/corbeillab/Ray-Cloud-Browser) is broken.
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