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Forum: RNA Sequencing 11-04-2018, 04:57 PM
Replies: 1
Views: 300
Posted By GenoMax
If you are using a specific kit then follow the...

If you are using a specific kit then follow the directions included with the kit for processing of data.
Forum: Illumina/Solexa 11-02-2018, 04:27 AM
Replies: 6
Views: 770
Posted By GenoMax
MiSeq is the champ of sequencing difficult...

MiSeq is the champ of sequencing difficult libraries. I would not compare success on a MiSeq to a different sequencer for odd libraries. If you are running an un-supported application then you are on...
Forum: Bioinformatics 10-29-2018, 04:57 AM
Replies: 657
Views: 137,687
Posted By GenoMax
@1989sn1027: Brian has not been participating on...

@1989sn1027: Brian has not been participating on SA for last few months. You could try to create a ticket (https://sourceforge.net/p/bbmap/tickets/)at Source Forge and see if he responds to this...
Forum: Bioinformatics 10-25-2018, 04:02 AM
Replies: 657
Views: 137,687
Posted By GenoMax
I see that you are assigning 60G of RAM. Have you...

I see that you are assigning 60G of RAM. Have you tried to assign more and see if it helps?
Forum: General 10-24-2018, 10:04 AM
Replies: 1
Views: 306
Posted By GenoMax
You could use bbsplit.sh from BBMap to figure...

You could use bbsplit.sh from BBMap to figure this out in one step. Take a look at this thread (http://seqanswers.com/forums/showthread.php?t=41288)about how to do this.

If you already have the...
Forum: RNA Sequencing 10-17-2018, 09:10 AM
Replies: 5
Views: 710
Posted By GenoMax
Not likely. For a sequencer it is just DNA. Does...

Not likely. For a sequencer it is just DNA. Does not matter if it came from rRNA or some other gene. If the two sets of runs happened on two different chemistries or different machines then may be...
Forum: Illumina/Solexa 10-12-2018, 07:24 AM
Replies: 2
Views: 525
Posted By GenoMax
This data must have been sequenced on NextSeq (or...

This data must have been sequenced on NextSeq (or NovaSeq). That string of "G" is probably no-signal which is interpreted as G base.

The bias at the beginning of the reads is likely due to the...
Forum: Illumina/Solexa 10-09-2018, 09:29 AM
Replies: 2
Views: 366
Posted By GenoMax
1. Yes (AFAIK). 2. Quality will be lower for...

1. Yes (AFAIK).
2. Quality will be lower for the last set of cycles. Depending on your library quality (and nucloetide diversity) that effect may be more or less pronounced.
3. Index reads are...
Forum: Bioinformatics 10-04-2018, 05:41 AM
Replies: 308
Views: 85,879
Posted By GenoMax
@FlySquirrelFly: It has become difficult to get a...

@FlySquirrelFly: It has become difficult to get a hold of Brian (due to his day job responsibilities) but I will flag your post for him to see if he can respond.

I recall from some past discussion...
Forum: Bioinformatics 09-27-2018, 06:31 PM
Replies: 2
Views: 374
Posted By GenoMax
Search EBI-ENA for minION. Here is a...

Search EBI-ENA for minION. Here is a representative search (https://www.ebi.ac.uk/ena/data/search?query=minion%2C+bacteria). Click on categories on the left (experiment is a good one).
Forum: Bioinformatics 09-27-2018, 05:23 AM
Replies: 4
Views: 374
Posted By GenoMax
This may be easier to do by obtaining the index...

This may be easier to do by obtaining the index read as a separate fastq file. You will also get real Q-scores for bases in index read when you do that. Stand-alone bcl2fastq allows one to get data...
Forum: Bioinformatics 09-26-2018, 12:01 PM
Replies: 4
Views: 374
Posted By GenoMax
Are you sure about that? Generally in case of...

Are you sure about that? Generally in case of Illumina sequencing index sequences (which are not part of actual reads) are transferred to the fastq header as a part of demultiplexing process. That is...
Forum: Bioinformatics 09-26-2018, 04:07 AM
Replies: 1
Views: 177
Posted By GenoMax
I am not sure what the size of BAM file you are...

I am not sure what the size of BAM file you are trying to use is but you could try adding the option "--java-mem-size=16G" to your qualimap command (Adjust RAM amount as needed). and see if that...
Forum: Illumina/Solexa 09-24-2018, 07:00 AM
Replies: 7
Views: 652
Posted By GenoMax
While likely let us hope that is not the case...

While likely let us hope that is not the case because otherwise you would be losing a lot of data to adapters and would have short reads.

When you have had a chance to investigate let us know...
Forum: Illumina/Solexa 09-24-2018, 06:11 AM
Replies: 7
Views: 652
Posted By GenoMax
Low nucleotide diversity in this case means...

Low nucleotide diversity in this case means majority of clusters will have e.g. an "A". When that happens the ability of the image analysis software to distinguish among clusters is hampered which...
Forum: Illumina/Solexa 09-24-2018, 04:20 AM
Replies: 7
Views: 652
Posted By GenoMax
1. Are these amplicons or sequences where there...

1. Are these amplicons or sequences where there is low nucleotide diversity expected?
2. You possibly have short inserts. Once you go through your inserts you are then sequencing into the adapter on...
Forum: RNA Sequencing 09-20-2018, 05:22 AM
Replies: 1
Views: 530
Posted By GenoMax
featureCounts is going to count reads per feature...

featureCounts is going to count reads per feature ("-t exon") that you specify and then will summarize the counts per gene ("-g gene_id"). It is going to report raw read counts and does not normalize...
Forum: Bioinformatics 09-09-2018, 08:30 AM
Replies: 1
Views: 459
Posted By GenoMax
I would suggest that you use the Illumina...

I would suggest that you use the Illumina provided adapter sequence. BBMerge detection feature is good when you don't have that information a priori. There may be some sequencing errors in your reads...
Forum: Illumina/Solexa 09-02-2018, 05:53 AM
Replies: 2
Views: 650
Posted By GenoMax
You just have to live with the quality of the...

You just have to live with the quality of the reads you get. Illumina requires 25 cycles in first read to accurately calibrate Q scores. Your type of sequencing is common with "Drop-seq" like methods...
Forum: Illumina/Solexa 08-21-2018, 12:24 PM
Replies: 5
Views: 896
Posted By GenoMax
If you are referring to sequences in...

If you are referring to sequences in "undetermined" files then these reads contain indexes that don't fit expected results (based on the samplesheet that you provide that has SampleID_Index mapping)....
Forum: Bioinformatics 08-14-2018, 04:21 AM
Replies: 3
Views: 699
Posted By GenoMax
You can try running the job again while nothing...

You can try running the job again while nothing else is running on the machine. You may be able to just get away with it.
Forum: Bioinformatics 08-13-2018, 12:02 PM
Replies: 3
Views: 699
Posted By GenoMax
Does 4TB refer to disk space? bwa is looking for...

Does 4TB refer to disk space? bwa is looking for more RAM so my guess is you don't have 53G of free RAM available or you are not the only user on this server.
Forum: Pacific Biosciences 08-12-2018, 04:29 PM
Replies: 4
Views: 1,531
Posted By GenoMax
Appears to some kind of file permission issue....

Appears to some kind of file permission issue. You should report this to PacBio tech support directly.
Forum: Bioinformatics 08-12-2018, 04:27 PM
Replies: 26
Views: 7,812
Posted By GenoMax
"bbsplit.sh" is a general purpose tool that will...

"bbsplit.sh" is a general purpose tool that will bin reads into any number of bins (depending on the reference sequences provided, you can provide as many as you want). In this case you would provide...
Forum: Bioinformatics 08-12-2018, 06:01 AM
Replies: 26
Views: 7,812
Posted By GenoMax
Have you tried using "bbsplit.sh" with human...

Have you tried using "bbsplit.sh" with human genome to see if that works better. If you are interested in non-human data then I would use the non-masked genome and risk losing a few additional reads.
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