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Forum: Sample Prep / Library Generation 07-31-2018, 12:55 PM
Replies: 2
Views: 898
Posted By fanli
iGenomX?

Anyone have experience/thoughts on the iGenomX Riptide system? Worth checking out..?
Forum: Bioinformatics 11-30-2017, 10:10 AM
Replies: 3
Views: 754
Posted By fanli
QIIME's split_libraries_fastq.py...

QIIME's split_libraries_fastq.py (http://qiime.org/scripts/split_libraries_fastq.html) script can probably do what you want. You can pretty easily create an index FASTQ file of your 10nt barcodes, if...
Forum: Illumina/Solexa 07-14-2017, 03:59 PM
Replies: 3
Views: 1,951
Posted By fanli
Maybe try looking in the index read files and see...

Maybe try looking in the index read files and see if the 8-base barcodes are indeed at the beginning of the index reads. --create-fastq-for-index-reads


Seems appropriate to me.
Forum: Bioinformatics 07-14-2017, 03:52 PM
Replies: 2
Views: 549
Posted By fanli
RNAstructure is another alternative: ...

RNAstructure is another alternative:
http://rna.urmc.rochester.edu/RNAstructure.html

CentroidFold is another:
https://github.com/satoken/centroid-rna-package

If you have that many sequences,...
Forum: Bioinformatics 07-14-2017, 03:46 PM
Replies: 26
Views: 7,805
Posted By fanli
I would caution against blindly filtering your...

I would caution against blindly filtering your metagenomic data against any database of contaminants. Ideally, you would have negative controls run alongside your samples that could be checked for...
Forum: Illumina/Solexa 07-10-2017, 11:09 AM
Replies: 6
Views: 981
Posted By fanli
I'm curious about this too - we've held back from...

I'm curious about this too - we've held back from running different amplicons together precisely because it'd be a complete shot in the dark as to how much to tweak loading concentrations.

Here is...
Forum: Bioinformatics 05-19-2017, 03:53 PM
Replies: 12
Views: 1,421
Posted By fanli
Have you tried contacting a university ombudsman...

Have you tried contacting a university ombudsman or someone similarly involved in research ethics? They may side with your collaborators, given that they represent the university but it may be...
Forum: Bioinformatics 05-19-2017, 12:55 PM
Replies: 12
Views: 1,421
Posted By fanli
Seems to me this is something you should have...

Seems to me this is something you should have taken up with your collaborators.

I would hesitate to call this malpractice, as the word has strong implications particularly in the medical field.
Forum: Bioinformatics 04-11-2017, 10:43 AM
Replies: 1
Views: 708
Posted By fanli
You can download either MFold...

You can download either MFold (http://unafold.rna.albany.edu/?q=mfold/download-mfold) or RNAfold (http://www.tbi.univie.ac.at/RNA/) as standalone software to do a batch.
Forum: Illumina/Solexa 04-11-2017, 09:33 AM
Replies: 2
Views: 1,003
Posted By fanli
I would tend to agree with @thermophile that...

I would tend to agree with @thermophile that custom primers would be the way to go here. If you are on a limited budget, Illumina kits tend to be quite expensive on a per-sample basis.

If you are...
Forum: Metagenomics 04-11-2017, 09:27 AM
Replies: 3
Views: 1,380
Posted By fanli
What amplicon are you using? Some of them have...

What amplicon are you using? Some of them have substantial variation in length in the variable region.

See:
https://www.biostars.org/p/78179/
Forum: Introductions 04-07-2017, 07:34 AM
Replies: 6
Views: 1,076
Posted By fanli
Another option: ...

Another option:
http://www.sequin.xyz/about/rnaseq/

In my experience, ERCC spikes aren't very useful for normalization. See this paper:
http://www.nature.com/nbt/journal/v32/n9/full/nbt.2931.html
Forum: Illumina/Solexa 03-23-2017, 10:03 PM
Replies: 29
Views: 6,224
Posted By fanli
@nucacidhunter, was this an identical pool in...

@nucacidhunter, was this an identical pool in terms of bacterial biomass and complexity? We've seen variation in sequencing quality for some amplicon libraries that probably had more off-target...
Forum: RNA Sequencing 03-13-2017, 01:35 PM
Replies: 7
Views: 1,603
Posted By fanli
I don't see anything in the traces that would...

I don't see anything in the traces that would make me overly alarmed. Most DGE software packages (edgeR, DESeq2, etc.) allow you to specify batch variables.
Forum: Illumina/Solexa 03-10-2017, 09:11 AM
Replies: 6
Views: 1,515
Posted By fanli
I'll just chip in my (rather long) 2c, as we've...

I'll just chip in my (rather long) 2c, as we've spent a great deal of time recently dealing with these issues.

First off, like @thermophile said, you absolutely need to include negative controls...
Forum: Bioinformatics 03-08-2017, 08:49 AM
Replies: 21
Views: 3,380
Posted By fanli
You might find this benchmark to be helpful: ...

You might find this benchmark to be helpful:
http://www.nature.com/articles/srep19233

My understanding is that you are better off using newer k-mer based approaches as opposed to BLAST. I've had...
Forum: Bioinformatics 03-08-2017, 08:39 AM
Replies: 21
Views: 3,380
Posted By fanli
Would something like kraken or CLARK not be...

Would something like kraken or CLARK not be helpful? Are you trying to assemble and annotate de novo genomes? Or trying to figure out the microbial composition and functional content? I guess my...
Forum: Bioinformatics 03-08-2017, 08:33 AM
Replies: 21
Views: 3,380
Posted By fanli
Out of curiosity, why are you joining the read...

Out of curiosity, why are you joining the read pairs? A lot of the metagenomics software out there now supports paired end reads as input. The metaSPAdes assembler @GenoMax mentioned requires paired...
Forum: RNA Sequencing 02-23-2017, 08:16 AM
Replies: 8
Views: 1,773
Posted By fanli
Perhaps you have mixed up files in your script....

Perhaps you have mixed up files in your script. You may want to check the logs in your tophat output directory.

As an example, here's what my align_summary.txt looks like:
Left reads:
...
Forum: RNA Sequencing 02-22-2017, 10:17 AM
Replies: 8
Views: 1,773
Posted By fanli
Like the prep_reads.info is from one sample and...

Like the prep_reads.info is from one sample and the tophat align_summary is from another?
Forum: RNA Sequencing 02-22-2017, 08:12 AM
Replies: 8
Views: 1,773
Posted By fanli
This says your input to tophat is only ~460k read...

This says your input to tophat is only ~460k read pairs. This directly contradicts what you posted in the prep_reads.info. Are you sure you don't have mismatched files?
Forum: Illumina/Solexa 02-22-2017, 08:03 AM
Replies: 29
Views: 6,224
Posted By fanli
We did a 2x300 run for a TCR library recently....

We did a 2x300 run for a TCR library recently. Passed spec (77.5% Q30 @ 939k with 15% PhiX), but clear tailing towards the ends of both R1 and R2 as well.

I could see error rates approaching 5%...
Forum: Bioinformatics 02-16-2017, 09:37 AM
Replies: 1
Views: 683
Posted By fanli
I think there is a decent amount of evidence out...

I think there is a decent amount of evidence out there that cuffdiff is less than optimal for gene-level DE analysis.

See:...
Forum: Bioinformatics 02-16-2017, 09:29 AM
Replies: 2
Views: 944
Posted By fanli
I like the phyloseq R package: ...

I like the phyloseq R package:
https://joey711.github.io/phyloseq/

If you are new to metagenomics work, you may want to start here:
http://qiime.org/
Forum: Epigenetics 12-21-2016, 10:10 AM
Replies: 3
Views: 3,986
Posted By fanli
Yeah, we've done some differential ChIP analysis...

Yeah, we've done some differential ChIP analysis but it was also pretty much seat-of-the-pants. We used a combination of your two approaches:
1) define a set of binding intervals based on a peak...
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