Tag: bcl2fastq

Extra-hematopoietic immunomodulatory role of the guanine-exchange factor DOCK2

Cell isolation, reprogramming and culture Approval was obtained for human cell and tissue sample collection and genetic reprogramming from the Institutional Review Board (protocols 19–252, 18–243, 21–060, 19–284 and 415-E/1776/4-2014, Ethics Committee of the province of Salzburg). Adult samples were collected in accordance with the Declaration of Helsinki after written…

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BCL compression

BCL compression 2 Hi, I’m trying to store some RUNs generated by different Illumina sequencers. My idea is to compress them to keep all files together (and reduce size, but this is secondary)… however, because the big amount of files and because the global size can be enormous (~120GB!) I’m…

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How do I get separate ADT / CITE-seq fastq’s from single SRA / BAM files? (originally generated from cellranger)

How do I get separate ADT / CITE-seq fastq’s from single SRA / BAM files? (originally generated from cellranger) 0 Hello all. I am trying to pre-process some single cell RNA and ADT (Totalseq-C) data from an GEO SRA, but having some issues getting separate fastq’s for the “CITE-seq” (ADT)…

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DRAGEN .bcl conversion error due to improper custom p5 oligos

DRAGEN .bcl conversion error due to improper custom p5 oligos 0 I am working on a custom library preparation method and I designed my p5 oligos incorrectly. To be specific, I used the reverse complement of the correct p5 index sequence. As a result my fastq files aren’t demultiplexing properly….

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Live-seq enables temporal transcriptomic recording of single cells

Biological materials RAW264.7, 293T and HeLa cells were obtained from ATCC. RAW264.7 cells with Tnf-mCherry reporter and relA-GFP fusion protein (RAW-G9 clone) were kindly provided by I.D.C. Fraser (National Institutes of Health). The IBA cell line derived from the stromal vascular fraction of interscapular brown adipose tissue of young male…

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bgzf_read_block] EOF marker is absent reformat.sh

BBMap/BBTools reformat.sh : real error or spurious message? [W::bgzf_read_block] EOF marker is absent reformat.sh 1 When subsampling paired-end .fastq.gz files using reformat.sh from BBMap/BBTools, I get this error message: [W::bgzf_read_block] EOF marker is absent reformat.sh I’ve checked the input files with gunzip -t, no error. The input files are a…

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BTG2 gene predicts poor outcome in PT-DLBCL

Introduction Primary testicular diffuse large B-cell lymphoma (PT-DLBCL) is a rare and aggressive form of mature B-cell lymphoma.1–3 PT-DLBCL was the most common type of testicular tumor in men aged over 60 and characterized by painless uni- or bilateral testicular masses with infrequent constitutional symptoms.4–6 PT-DLBCL shows significant extranodal tropism,…

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Extracellular circulating miRNAs as stress-related signature to search and rescue dogs

Study approval was provided by the Research Ethics Committee of the University of Perugia (report n.2018-21 of 11/12/2018) according to Italian Ministry of Health legislation18. All methods were carried out following relevant guidelines and regulations and the study was carried out in compliance with the ARRIVE guidelines. Informed consent is…

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Merging compressed fastq files based on a conditions defined in a csv file

Hello everybody, I have a question quite different about similar topic addressed on: Post not found I tried Paul’s bash script in the web indicated above (fastq_lane_merging.sh) adapting to my filename organization data being: #!/bin/bash for i in $(find ./ -type f -name “*.fastq.gz” | while read F; do basename…

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empty fastq files created by docker bcl2fastq2 v2.20 OSX

empty fastq files created by docker bcl2fastq2 v2.20 OSX 0 HI, I installed a following docker image, REPOSITORY TAG IMAGE ID CREATED SIZE zymoresearch/bcl2fastq latest 037f216c2523 13 months ago 117MB and run a following command; docker run -d –name bcl2fastq -v /Volumes/Aura2/bcl_NU/170720_NB501488_0132_AH5V32BGX3:/mnt/run -v /Volumes/Aura2/output:/mnt/out zymoresearch/bcl2fastq:2.20 -R /mnt/run -o /mnt/out/Data/Intensities/BaseCalls/Alignment_1 –barcode-mismatches…

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Illumina summary.csv file units

Illumina summary.csv file units 1 We are using illumina Novaseq machines to do some sequencing and one of the files produced is a summary.csv file that has some info about the run that was sequenced. However, it is tough to figure out what units were used for a lot of…

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bcl2fastq with xGen Dual Index UMI Adapters to produce 3 read and 2 index fastqs

I usually remove Ns from the sample sheet and then run this command $ bcl2fastq -i Runfolder/Data/Intensities/BaseCalls -R Runfolder -o output_directory –sample-sheet Runfolder/SampleSheet.csv -r number_of_threads -p number_of_threads -w number_of_threads –no-lane-splitting –mask-short-adapter-reads 0 –use-bases-mask Y*,I8Y9,I8,Y* Original sample sheet: Sample_ID,Sample_Name,Description,index,I7_Index_ID,index2,I5_Index_ID,Sample_Project Sample1,Sample1,,TACCGAGGNNNNNNNNN,TACCGAGGNNNNNNNNN,AGTTCAGG,AGTTCAGG,BRCA1 Sample2,Sample1,,CGTTAGAANNNNNNNNN,CGTTAGAANNNNNNNNN,GACCTGAA,GACCTGAA,BRCA1 Formatted sample sheet: Sample_ID,Sample_Name,Description,index,I7_Index_ID,index2,I5_Index_ID,Sample_Project Sample1,Sample1,,TACCGAGG,TACCGAGG,AGTTCAGG,AGTTCAGG,BRCA1 Sample2,Sample1,,CGTTAGAA,CGTTAGAA,GACCTGAA,GACCTGAA,BRCA1 Our RunInfo.xml file looks…

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BCL files conversion to FASTQ without SampleSheet.csv

BCL files conversion to FASTQ without SampleSheet.csv 1 Dear community, I have got NGS data which is basically the BaseCalls folder with .bcl files. I want to know how to successfully convert .bcl files to .fastq format. So far, I have been using the bcl2fastq program, however, I have no…

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Why is bcl2fastq2 taking so long to calculate stats?

Our lab has been using bcl2fastq v2.20.0.422 to demultiplex RNA-seq data sequenced on an Illumina Novaseq machine on a beefy EC2 instance and we’ve run into the strange problem: namely that while demultiplexing is very fast, generating the stats files are unbearably slow. In a recent example, we demultiplexed one…

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Troubleshooting Tips – bcl2fastq creates duplicate reads

Forum:Troubleshooting Tips – bcl2fastq creates duplicate reads 1 Hi, I have seen a few times where bcl2fastq (v2.20) will produce duplicate FASTQ entries in sequencing read IDs, raw sequences, & quality scores. This causes issues with downstreams tools like Picard MarkDuplicates (e.g. Exception in thread “main” htsjdk.samtools.SAMException: Value was put…

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Get failed reads on Novaseq

Get failed reads on Novaseq 0 Dear all, on a NextSeq550, I can get failed reads (not PF) with –with-failed-reads using bcl2fastq. With the more modern bclconvert software, this does not exist. support.illumina.com/bulletins/2020/10/upgrading-from-bcl2fastq-to-bcl-convert.html Also, it seems failed reads are not saved any more by the Novaseq, which is problematic for…

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How to fasterq-dump 10x genomics snATACseq fastq from SRA

How to fasterq-dump 10x genomics snATACseq fastq from SRA 2 I am trying to retrieve fastq files from a 10x genomics snATACseq dataset on SRA. Each run should have 4 fastq files associated with it: I1: Dual index i7 read (optional) R1: Read 1 R2: Dual index i5 read R3:…

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bcl2fastq ERROR

bcl2fastq ERROR 1 Hi all, I have a problem. I’m running bcl2fastq software on Ubuntu, but I can’t demultiplex anything as the software gives me this error: *ERROR: bcl2fastq::common::Exception: 2021-Sep-02 12:12:34: Success (0): /home/Software/bcl2fastq/src/cxx/lib/layout/Layout.cpp(693): Throw in function void bcl2fastq::layout::handleDefaultSample(bool, bool, size_t, bcl2fastq::layout::LaneInfo&, bool&) Dynamic exception type: boost::exception_detail::clone_impl<bcl2fastq::common::InputDataError> std::exception::what: Missing a…

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