Tag: FAST5

The landscape of genomic structural variation in Indigenous Australians

Cohorts Saliva and/or blood samples were collected from consenting individuals among four NCIG-partnered communities: Tiwi Islands (comprising the Wurrumiyanga, Pirlangimpi and Millikapiti communities), Galiwin’ku, Titjikala and Yarrabah, between 2015 and 2019. Non-Indigenous comparison data, generated from unrelated Australian individuals of European ancestry, was drawn from two existing biomedical research cohorts:…

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Resin acids play key roles in shaping microbial communities during degradation of spruce bark

Bark preparation Spruce bark was obtained from the Iggesund pulp and paper mill (Iggesund, Holmen AB, Sweden), from a bark pile resulting from stripping of spruce logs at the mill after harvest, with the average age of trees at harvest being ~70 years. The bark was left to dry at…

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Generating high-quality plant and fish reference genomes from field-collected specimens by optimizing preservation

Sample collection A total of nine species of marine fish were collected across three different sampling days (September 7th, 9th, and 12th 2022) under IACUC Animal Use Protocol S12219 (Supplementary Data 1). Six species were collected using a speargun donated by a local fisher. Fish were transported back to shore, euthanized,…

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Common analysis of direct RNA sequencinG CUrrently leads to misidentification of m5C at GCU motifs

Introduction Oxford Nanopore Technologies (ONT) direct RNA sequencing (Fig 1A) enables detection of RNA modifications. A modified base produces an altered electrical current and/or dwell time relative to a canonical base that can be detected with algorithms (Garalde et al, 2018; Smith et al, 2019; Workman et al, 2019). Figure…

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Visualize and explore eventalign data against reference

Visualize and explore eventalign data against reference 0 Hi all, Is anyone aware of a tool (GUI or python/R/bash package) to explore eventalign nanopore data (or fast5 raw data) with the corresponding alignment to a reference genome? Kind of like viewing how reads in a bam file align against a…

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The Biostar Herald for Monday, November 20, 2023

The Biostar Herald publishes user submitted links of bioinformatics relevance. It aims to provide a summary of interesting and relevant information you may have missed. You too can submit links here. This edition of the Herald was brought to you by contribution from Istvan Albert, and was edited by Istvan…

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Base modification detection for direct RNA sequencing Nanopore

Base modification detection for direct RNA sequencing Nanopore 0 We have Nanopore RNA sequencing data (fast5) generating using the RNA002 chemistry. We are interested in determining the methylation status (5mc) of these reads. At least initially, we don’t even need to align the reads, we just want to know which…

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Apply to install Guppy

Apply to install Guppy 0 Hi dear community I really need a GPU version of Guppy(>=3.6.1 version), but I’m not eligible to download Guppy. Could someone who has the ability to download this tool please send a copy to my email?(rogierzhang@126.com) Thank you very much. community.nanoporetech.com/downloads Guppy files fast5 ONT…

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Does Guppy have no –fast_out parameters?

Does Guppy have no –fast_out parameters? 0 Dear whom it may concern: I want to obtain the 6mA methylation modification of the genome through the fast5 file of ONT sequencing. The first thing I needed to fix was that the fast5 file from the sequencing company doesn’t have the events…

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Porechop demulitplexing of nanopore sequencing data and barcode bins for each sample

Porechop demulitplexing of nanopore sequencing data and barcode bins for each sample 0 Dear, I am new to nanopore community and have some questions. I might sound silly! We preapred nanopore librariy using 1-24 SQK-16S024 kit for 24 samples. After sequencing, i got fast5 and fastq files. Fastq files were…

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Ultra-fast deep-learned CNS tumour classification during surgery

Data simulation Short nanopore sequencing runs yield sparse and random coverage of the genome. To enable model training, we generate simulated sparse nanopore runs based on microarray data. To this end, N simulated reads are randomly sampled from the read length distribution (D) and assigned a start mapping position in…

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Scaling logical density of DNA storage with enzymatically-ligated composite motifs

Composite motifs as building blocks for DNA storage A composite motif is a representation of a position in an oligo sequence that uses a combination of motifs drawn from a fixed motif library to encode data. For example, assuming a library of 32 motifs, and a combination factor of four,…

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mRNA vaccine quality analysis using RNA sequencing

Design and synthesis of reference plasmid A reference construct was first designed, with the intention of optimising the production of RNA therapeutics for pre-clinical research. The coding sequence of eGFP30 was selected as a reporter in the coding region, as its protein product can be assayed simply through Flow cytometry…

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FastQC – Browse /v0.12.0 at SourceForge.net

RELEASE NOTES FOR FastQC v0.12.0 This release introduces new functionality as well as resolving some existing problems. Fix bugs in the per tile plot if zero length sequences are present Add a count of total bases to the Basic Stats output Use better sorting of the best contaminant finding Add…

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Cannot process all the reads in a fast5 file?

Cannot process all the reads in a fast5 file? 0 Hello all, I have a ~296GB .fast5 which is the result of a metagenomic effort. I don’t have a precise understanding of the history of this file, but I believe it is the product of multiple FAST5 being concatenated. Others…

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CUDA error when I run megalodon (2.4.1) and guppy (6.0.1+652ffd1) with GPU support.

CUDA error when I run megalodon (2.4.1) and guppy (6.0.1+652ffd1) with GPU support. 1 Hello, everyone, recently when I run the megalodon command, there was a constant error. attached is my demo in the script. megalodon \ ~/01.data/01.ONT_data/02.ONT_test/AW/FAST5_PASS \ –guppy-params “-d /public/home/zenglingsen/04.software/03.Guppy/rerio/basecall_models/” \ –guppy-server-path /public/home/zenglingsen/04.software/03.Guppy/ont-guppy/bin/guppy_basecall_server \ –guppy-config res_dna_r941_prom_modbases_5mC_CpG_v001.cfg \ –outputs…

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Processing Fast5 files from ONT for direct RNA

Processing Fast5 files from ONT for direct RNA 2 Hi, Am trying to process some sequencing data from ONT. I have fast5, fastq, fasta files. I have also generated bam files with minimap2 and samtools. However, when I cam trying to run Nanopolish, ELIGOS2, DRUMMER, Tombo – all tools are…

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Universal whole-genome Oxford nanopore sequencing of SARS-CoV-2 using tiled amplicons

Clinical RNA SARS-CoV-2 isolate For surveillance studies, a set of residual nasopharyngeal swab specimens positive for SARS-CoV-2 qRT-PCR from the Republican Diagnostic Centre (RDC) (umc.org.kz/en/) and private laboratory KDL “Olymp” (www.kdlolymp.kz/) were collected across the Republic of Kazakhstan between 2020 and 2022. 341 sequences that passed quality control are deposited…

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guppy basecaller giving error while basecalling

guppy basecaller giving error while basecalling 0 I am trying to basecall the nanopore fast5 file using the guppy basecaller version 6.4.6 (GPU). Its giving me the following error: [guppy/error] main: CUDA error at /builds/ofan/ont_core_cpp/ont_core/common/cuda_common.cpp:182: [guppy/warning] main: An error has occurred. Aborting. Can someone please help how to solve this…

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Custom barcodes on guppy

Custom barcodes on guppy 0 Hello everyone, I’m intending to use guppy to read fast5 files generated by a Nanopore PromethION, but I’m planning to use custom barcodes (not the 96 barcodes kit from Nanopore). My questions are: Is guppy neural network trained to only identify these 96 barcodes, or…

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Y chromosome sequence and epigenomic reconstruction across human populations

Data production Complete Y chromosomes from six different human haplogroups were isolated as previously described17 (Fig. 1a, b). In brief, chromosomes were obtained from lymphoblastoid cell lines (LCLs) used in the 1000 Genomes Project31 (1kgp) and sequenced on the ONT MinION. We also made use of the Y chromosome sorted ONT…

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Reads with highest MAPQ values from SAM files are showing mismatches to reference sequence and IGV classified them as supplementary reads

Hi all, I am expressing a GFP synonymous variant library in human cells and sequencing its RNA on the nanopore and I am having some trouble analysing the data. Initially, I basecalled all the fast5 files using the super accuracy model in the guppy basecaller, then I discarded the reads…

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Convert Nanopore Fast5 files to Fasta format

Convert Nanopore Fast5 files to Fasta format 0 Hi, Could anybody please suggest me a tool to convert the Nano-pore fast5 files to fasta files ? I have already tried Poretools and Nanopolish but wasn’t successful. I understand that fast5 can be converted to fastq directly but I need them…

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Problem with GPU guppy_basecaller and SLURM

Hello, I am having trouble running the GPU version of guppy on a cluster using SLURM. I have two guppy scripts called guppy_pass1.sh and guppy_pass2.sh. In the pass 1 script I basecall to get look at the ‘raw’ data without additional filtering/trimming. In the pass 2 script I basecall to…

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Babraham Bioinformatics – FastQC A Quality Control tool for High Throughput Sequence Data

FastQC Function AMPERE quality control tool for elevated throughput sequence data. Your Java What A match Java Runtime Ecology This Picard BAM/SAM Libraries (included in download) Code Maturation Robust. Mature code, but feedback exists comprehended. Code Released No, under GPL v3 or later. Initial Contact Simon Andrews Download Now Views…

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Gut microbiome profiling of neonates using Nanopore MinION and Illumina MiSeq sequencing

Introduction Perturbations in the infant gut microbiome during early life affect growth, development, and long-term health (Sherman, 2010; Carl et al., 2014; Tarr and Warner, 2016). Preterm infants have a physiologically and anatomically immature gastrointestinal tract that is known to be more permeable than that of term infants, and their…

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Best tool for trimming and filtering nanopore direct RNA sequencing.

Best tool for trimming and filtering nanopore direct RNA sequencing. 1 Hello, I am new to analyzing long read data. I am trying to create a pipeline for analyzing data from Oxford Nanopore’s MinION Mk1b using the R.9.4.1 flow cell and using the Direct RNA Sequencing Kit. We used only…

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Why does Guppy output different sequence data (same model)?

I ran a nanopore sequencing on mk1c device with live basecalling and obtained some fastq in fastq_pass, fastq_fail folders. I tried to rerun the basecalling in a different machine, but found that they produce different sequences in just a test case, e.g.: In the fastq basecalled in mk1c > @b2e79451-050f-4d74-b091-40e6b6ee2229…

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TUT4/7-mediated uridylation of a coronavirus subgenomic RNAs delays viral replication

Cell lines and tissue culture The murine 17-CL1 cell line (derived from 3T3 cells, BEI Resources, Cat. No. NR-53719) was maintained in Dulbecco Modified Eagle Medium (DMEM, Life Technologies) supplemented with 10% Fetal Bovine Serum (FBS, Gemini Bio-Products), L-glutamine (Invitrogen), and Sodium pyruvate (Invitrogen). The NCTC clone 1469 cell line…

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How to rerun Guppy Basecalling (with both fast5_pass and fast5_fail)

How to rerun Guppy Basecalling (with both fast5_pass and fast5_fail) 0 Hi, I recently started a nanopore sequencing and used the live-basecalling feature in mk1c. It was proceeding very slowly. I want to free up the machine to do more sequencing and am thinking of recovering the raw data in…

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Evaluation of Mycobacterium tuberculosis enrichment in metagenomic samples using ONT adaptive sequencing and amplicon sequencing for identification and variant calling

Enrichment in ONT amplicon sequencing and adaptive sequencing The number of TB reads detected in clinical samples can vary from one to thousands, with over 90% human reads8,9. To simulate a simple TB metagenomic sample for enrichment evaluation using MinION amplicon sequencing and adaptive sequencing, we prepared a synthetic metagenomic…

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How to remove duplicate reads after ONT basecalling from fastq.gz files

How to remove duplicate reads after ONT basecalling from fastq.gz files 1 Hi I ran into an error when my PC was halfway through basecalling, it errored on a fast5 file – number 577. I restarted basecalling from 578, and then when finished I basecalled 577 on its own. It…

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demultiplexing with guppy_basecaller and guppy_barcoder using –detect_mid_strand_adapter –detect_mid_strand_barcodes produce > 90% of unclassified reads

A similar question has been already posted on the ONT community forum but without answers form guppy devs. I am using guppy 6.0.1 In short, when demultiplexing during basecalling using the options –do_read_splitting –detect_mid_strand_adapter –detect_mid_strand_barcodes ./guppy_basecaller -c dna_r9.4.1_450bps_sup.cfg -i $SCRATCH/barcode/fast5 -s $SCRATCH/barcode/fastq -x ‘auto’ –recursive –barcode_kits “EXP-NBD104” –trim_barcodes –trim_adapters –do_read_splitting…

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sequence alignment – Help with MinION sequencing data species identification

Hi I’m new to bioinformatics and have just completed my first run on the MinION (long read sequencing Oxford Nanopore Technologies). I was hoping someone could direct me towards R packages, workflow, tutorials or guides that will help me identify species that are present in my sample mainly for fungi…

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ONT analysis on Ubuntu

ONT analysis on Ubuntu 0 Hello guys, I am new on bioinformatic analysis of data from ONT (MinION with FLOW-MIN106D). The lab sent me a collection of fast5 files I have to analyze to obtain the metagenomic details (viruses). I have the new machine with on board Ubuntu 21 (Nvidia…

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MinION Data Examples (FAST5) Database

MinION Data Examples (FAST5) Database 0 Hello everyone, I am constructing a pipeline to analyze Oxford Nanopore MinION data. I have start from FAST5 files and for some optimizations I will try multiple tools for each step. So I will need several datasets to try. As I see most of…

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