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Forum: Sample Prep / Library Generation 10-02-2016, 02:11 AM
Replies: 13
Views: 2,585
Posted By avo
Adapting TruSeq PCR free for approx. 1000 bp...

Adapting TruSeq PCR free for approx. 1000 bp insert size is possible. The following plot is from a library size selected on a BluePippin after adapter ligation.

https://i.imgsafe.org/0cd6a5ad32.png
Forum: Illumina/Solexa 04-08-2016, 02:57 AM
Replies: 6
Views: 1,350
Posted By avo
Probably worth checking a few thumbnail images to...

Probably worth checking a few thumbnail images to have at least an estimate if and how many clusters are present.

I would assume either massive over- or underclustering
Forum: Sample Prep / Library Generation 08-17-2015, 01:21 AM
Replies: 5
Views: 4,863
Posted By avo
AMPure PB / AMPure XP / SPB differences

Does anyone know what the differences between the AMPure PacBio, AMPure XP or Illumina sample purification beads are? Different PEG concentrations,or if there no differences at all?

Thanks
Forum: Illumina/Solexa 11-12-2014, 03:56 AM
Replies: 4
Views: 3,587
Posted By avo
I agree with Brian. Are you sure it is a TruSeq2...

I agree with Brian. Are you sure it is a TruSeq2 library? We often see this kind of sequence content plots for Nextera libraries. In this case you should just use the NexteraPE-PE.fa adapter file.
Forum: Bioinformatics 10-20-2014, 11:35 PM
Replies: 17
Views: 7,520
Posted By avo
Can you give some more information about the run...

Can you give some more information about the run itself? Sample, Cluster density, Insert size, size selection and library prep might be helpful for troubleshooting.

It might be worth checking if...
Forum: RNA Sequencing 10-09-2014, 12:37 AM
Replies: 6
Views: 2,743
Posted By avo
Does Fastqc give you a warning for...

Does Fastqc give you a warning for overrepresented sequences?

I don't know the default -k setting for fastqc but maybe it helps to increase it.
My impression is that the adapter trimming worked....
Forum: Illumina/Solexa 09-26-2014, 05:10 AM
Replies: 27
Views: 8,285
Posted By avo
I'm a bit confused by the %Q30 graph shown here....

I'm a bit confused by the %Q30 graph shown here. I have seen 600V3 runs on our MiSeq that looked much worse. Yet we generally cluster well above 566k, so that might be a factor. Nevertheless I would...
Forum: De novo discovery 07-21-2014, 01:20 AM
Replies: 5
Views: 3,427
Posted By avo
In my experience the fastqc quality plots look...

In my experience the fastqc quality plots look similar to what we see with TruSeq libraries.
However i always do the trimming for adapters and quality.
Especially with Nextera, the bead size...
Forum: Bioinformatics 07-14-2014, 05:57 AM
Replies: 12
Views: 2,097
Posted By avo
you could also check Platanus...

you could also check Platanus (http://platanus.bio.titech.ac.jp/platanus-assembler/). It uses a multi-kmer approach and can handle kmer sizes > 127. We obtain good results with MiSeq 2x300bp reads.
Forum: Bioinformatics 06-11-2014, 03:58 AM
Replies: 4
Views: 3,827
Posted By avo
I would rather do this with the whole dataset. If...

I would rather do this with the whole dataset. If you have enough coverage (approx. >30-fold) the k-mer graph should not only be able to give you a hint about the genome size and coverage but also...
Forum: Sample Prep / Library Generation 05-26-2014, 06:00 AM
Replies: 5
Views: 4,029
Posted By avo
What kit did you use for the qPCR quantification...

What kit did you use for the qPCR quantification and how much library did you use for the first dilution? You could reduce the volume of the last elution step to 20 Ál. If thats not enough you can...
Forum: Illumina/Solexa 05-23-2014, 06:02 AM
Replies: 9
Views: 5,290
Posted By avo
Did any of these runs use the new MCS Version...

Did any of these runs use the new MCS Version 2.4.1.3?
To me it triggers an error that the number of cycles is out of range for the used cartridge.

Maybe the number of cycles is now limited in...
Forum: Illumina/Solexa 05-15-2014, 01:49 AM
Replies: 9
Views: 5,290
Posted By avo
Do you do those 350x300 or PE325 reads with the...

Do you do those 350x300 or PE325 reads with the standard 600-cycle kit, where the extra cycles derive from not reading indices and excessive reagents or is it some sort of "hacked" cartridge (like...
Forum: Sample Prep / Library Generation 11-15-2013, 01:57 AM
Replies: 2
Views: 2,238
Posted By avo
TruSeq PCR-free transition

Hi together,

my lab is planning to switch to the TruSeq PCR free library prep. We would like to perform size selection on a bluepippin. Reading various threads here about the low yields of the...
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