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Questions tagged [nanopore]

Questions specific to nanopore sequencing. For general question about long reads, use tag long-reads instead and for questions about specific sequencer use a specific sequencer tag (i.e. minion, gridion)

3
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1answer
41 views

Error correction within the long read

I am going to get some data from plasmid sequencing to identify SNPs on the plasmids. What is done in the lab is the following: The plasmids are purified by size. We amplify the plasmids using the ...
1
vote
1answer
33 views

Using ONT MinION, why is that we cannot get a full length DNA read?

Here, https://www.youtube.com/watch?v=E9-Rm5AoZGw, ONT claims they can read non-fragmented DNA and that with the hairpin like structure the double strands can be read at once. Then why is that MinION ...
0
votes
0answers
13 views

Is there a tool that converts fastq to fast5? [duplicate]

We need lots of fast5 files. Anyway i can convert fastq to fast 5 or simulate fast5?
5
votes
3answers
127 views

Total reads aligning to each reference within a bam file

I have two PCR amplicons that have been multiplexed and sequenced using the nanopore minion. I have aligned the fastq reads using minimap2 with a reference file containing both amplicon sequences and ...
3
votes
1answer
62 views

How to simulate nanopore reads?

I have looked already here: Tools for simulating Oxford Nanopore reads . This doesn't answer my question, because it lists a few Nanopore read simulators, but I have specific problems with each of ...
4
votes
1answer
178 views

Extracting strand specific reads from MinION cDNA-PCR protocol

I recently performed my first MinION run, and now I'm trying to analyze the data. Being pretty new to the bioinformatics field, I was hoping some of you could help me out. As a bit of background, I'...
3
votes
2answers
240 views

Can I stop my nanopore sequencing run if there are no more reads being produced?

I am sequencing a whole genome on MinION. I have used this flow cell for 6 hours for the phage lambda control. That gave me around 300,000 reads. I started a 48h sequencing run on the same flow cell ...
2
votes
0answers
52 views

aligner for 1D^2 oxford nanopore data

I am wondering if anyone has tried aligning 1d^2 RNA-seq data. I am trying that on minimap2. It is giving me very low ~50% of mapped reads. it gives ~95% mapped reads for 1D. I am using the following ...
3
votes
1answer
331 views

How many reads has my sequencing run produced on minion?

I am running a 48 h sequencing protocol on a FLO-MIN107, on MinION, with DNA library-prepped with SQK-RAD004. MinKNOW is installed on my Mac and controlling the MinION. I have found the following ...
2
votes
1answer
408 views

Albacore wheel not supported on this platform (Mac OS X El Capitan)

I download the Albacore wheel from Oxford Nanopore. I activated my conda environment that has Python 3 as the default python version. I tried to install Albacore with ...
1
vote
1answer
110 views

Basecalling process carried out by full_1dsq_basecaller.py/.exe

I am unable to understand what exactly the full_1dsq_basecaller.py/.exe script does. Is it giving the consensus sequence for linear and complementary strand, or it is just giving the reads that came ...
1
vote
1answer
708 views

Set directory where MinKNOW writes FAST5 files

I would like to set the folder in which MinKNOW writes the raw data. How can I do this? I do not currently know where MinKNOW will output my data. What is the default directory where MinKNOW outputs ...
5
votes
2answers
98 views

Can I use my computer while MinION is sequencing without negatively affecting the run?

I have a Mac Book Pro and I am about to start a sequencing run on the MinION. MinKNOW is going to be running for 2 days. Can I use my computer during sequencing? Can I browse the web and use Excel, ...
2
votes
1answer
722 views

Difference between 1D and 1D^2 data

I am working on ONT data. Initially, I have worked on data from 1D using Minimap2 aligner. I came across 1D^2 from ONT website. I was wondering how different is the data from two techniques? What is ...
1
vote
1answer
123 views

What to do with the configuration test cell after configuring the MinION with it?

What to do with the configuration test cell after configuring the MinION with it? I received a MinION with a configuration test cell in it. After running a configuration in the MinKNOW software while ...
3
votes
3answers
842 views

What are the pros and cons of the different basecallers in Oxford Nanopore Technology Sequencing?

What are the pros and cons of the different basecallers in Oxford Nanopore Technology Sequencing? I am about to start a MinION run on my laptop. What should I consider when choosing my basecaller? ...
3
votes
1answer
250 views

Does MinKNOW work with Mac OSX high sierra 10.13.1?

I have done a compatibility check as described here on my computer to know if I can use MinKNOW on my mac laptop. It says that my OS is incompatible and that the OSX must be Yosemite or El Capitan. ...
3
votes
1answer
176 views

Removing repeated reads from nanopore 1D² reads

This is summary of discussions and a question that was posted by Devon O'Rourke on Twitter Following Albacore basecalls on a 1D² library I get two sets of .fq files, summary stats, etc.: one for the ...
4
votes
3answers
185 views

sort a fasta file containing the Oxford Nanopore Technologies (ONT) header by sequencing start_time ascending

I have basecalled ONT reads and converted them to multifasta. The multifasta contains the original ONT headers in this format: ...
7
votes
2answers
126 views

Canu assembly not making a single consensus?

I've downloaded reads from this BioProject. Using canu with default parameters (no correction), I've got 4 contigs, none of which really look like the reference plasmid here. The command I used was: ...
4
votes
2answers
259 views

How to get Nanopore MinION fast5 from SRA

I found some Nanopore MinION data on SRA, which I would like to investigate. I use sratoolkit for Illumina data all the time, but I am not sure how to get the ...
5
votes
1answer
274 views

STAR-long parameters for aligning RNA ONT reads to genome

Are there any suggested parameters to align ONT reads to the reference genome using STAR-long? For now, I used the parameters suggested here, but I noticed a weird behaviour. I have RNA reads (D. ...
4
votes
2answers
153 views

Error rate setting in Canu error correction

I want to use Canu to correct my nanopore long read (version: MinION R9.5), but I am not quite sure how to set the correctErrorRate. Should I follow the Canu manual (Nanopore R7 2D and Nanopore R9 1D ...
7
votes
4answers
77 views

Introduce errors in reference transcripts according to external dataset error model

I would like to modify some reference transcripts from Ensembl (D. melanogaster) to introduce a controlled rate of random errors in the sequences. The idea would be to introduce random base ...
6
votes
1answer
131 views

Split FASTQ and matching BAM into matching chunks

I am running a slow downstream analysis on a large set of nanopore reads (approx 3 million), and would like to split them into smaller chunks, run the analysis in massively parallel, and then ...
9
votes
2answers
478 views

How to convert fastq to fast5

fast5 is a variant of HDF5 the native format in which raw data from Oxford Nanopore MinION are provided. You can easily extract ...
6
votes
3answers
284 views

Extract nanopore read ID & start times from fastq file

I have a fastq file from minION (albacore) that contains information on the read ID and the start time of the read. I want to extract these two bits of information into a single csv file. I've been ...
6
votes
1answer
554 views

How can I use Nanopore reads to close gaps or resolve repeats in a short-read assembly?

Low coverage MinION reads should be useful to close gaps and resolve repeats left by short-read assemblers. However, I haven't had any success with the software I know about. I'm aware of the ...
8
votes
2answers
645 views

How do you generate read-length vs read-quality plot for long-read sequencing data (e.g., MinION)?

How do you generate read-length vs read-quality plot (heat map with histograms in the margin) for long-read sequencing data from the Oxford Nanopore Technologies (ONT) MinION? The MinKNOW software ...
5
votes
2answers
169 views

How to estimate whether a long-read is meaningful sequence?

The setup Imagine that I work on an organism without a reference genome, and that the closest reference genome I can get is quite diverged. E.g. ~10% diverged in terms of SNVs when measured with ...
3
votes
2answers
308 views

Minion channel ID's from Albacore

The latest version of Albacore from Oxford Nanopore Technologies calls bases from raw fast5 files. A useful piece of output is the sequence_summary.txt, which is a ...
8
votes
3answers
203 views

Visualisation of long read RNA-Seq splicing

I have a dataset of Oxford Nanopore cDNA reads. Many of my reads are full-length or close to full-length transcripts, and I and am interested in examining alternative splicing. For this, I would like ...
7
votes
2answers
228 views

Building STAR Genome Index for nanopore RNA sequencing

I am aligning a dataset of 1,000,000 reads oh human mRNA sequenced on Oxford Nanopore Technologies' MinION, and would like to use the STAR aligner, using the parameters recommended by Pacific ...
8
votes
0answers
191 views

Improving consensus assembly from UMI-tagged nanopore reads

The Albertsen lab has recently put out a competition/challenge for read error correction I only found out about this today, and I think the conference where high-ranking participants were going to be ...
4
votes
1answer
115 views

Detecting structural variants with MinION data

Working on various cancers I have an interest in detecting structural variation (SV) in human, we've successfully used various tools like Pindel, SVDetect, Manta, and LUMPY, to name a few for ...
12
votes
2answers
79 views

Finding the location and unit length of repetitive sequences within a long read

After discovering a few difficulties with genome assembly, I've taken an interest in finding and categorising repetitive DNA sequences, such as this one from Nippostrongylus brasiliensis [each base is ...
4
votes
1answer
112 views

Rapid metagenomics classifiers on long read data [closed]

I recently used the minION (Nanopore, 9.4 flow cell, RAD001 kit) to generate a metagenome out of environmental samples. Passed reads weren't brilliant (196, average 1,594bp lenght), but working with ...
15
votes
1answer
313 views

How can I improve a long-read assembly with a repetitive genome?

I'm currently trying to assembly a genome from a rodent parasite, Nippostrongylus brasiliensis. This genome does have an existing reference genome, but it is highly fragmented. Here are some ...
15
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5answers
312 views

Tools for simulating Oxford Nanopore reads

Are there any free open source software tools available for simulating Oxford Nanopore reads?
22
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7answers
2k views

Read length distribution from FASTA file

I have a ~10GB FASTA file generated from an Oxford Nanopore Technologies' MinION run. How can I quickly and efficiently calculate the distribution of read lengths? A naive approach would be to read ...
13
votes
1answer
114 views

Compare alignment quality of multiple sequencing runs aligned against the same reference genome

I have run Oxford Nanopore Technologies' MinION sequencing on the same DNA sample using three flowcells, each aligned against the same reference genome (E.coli K12 MG1655) using both BWA MEM and ...