Questions tagged [assembly]

Process of creating the original sequence from the read sequences that it generated during a sequencing experiment. Can refer to genome assembly, in which case the original sequence is a genome, or transcripts assembly, in which case the original sequences are RNA transcripts.

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Can I get longer contigs by changing MEGAHIT settings?

My current settings are megahit -r input.fq --num-cpu-threads 32 --min-contig-len 300 --presets meta-large -o output. I picked the 'presets meta-large' because I am ...
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What assembler is appropriate for High-Fidelity PacBio reads

What assembler is appropriate for High-Fidelity PacBio reads? For example, canu is good for high-error PacBio reads. But what algorithm to use for HiFi reads? Would it be OK to use canu without the ...
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mapping and de novo assembly of plant without reference genome

I'm studying on a plant that has no reference genome and only has one scaffold assembly and one gff3 annotation file. Can I create an index with the same assembly and gff3 in STAR and do the mapping? ...
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nanopore - where to retrieve information from the basecaller used

Where can I find information about the basecaller used in a specific nanopore run (we have a Gridion machine)? I know already that we have Guppy v. 3.0.3, which I ...
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Finding simple sequence from reads with significant overlap

I wrote a script to pull out reads from a huge fastq file in an iterative manner, by finding homology to the previous sequence. It seems like it should be relatively easy to overlap them and ...
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fixing chromosome with new contig

I want to fix a chromosome of around 50-100Mb (chr.fasta) for a region that is missing a few kilobases of data. I have the missing sequence with a window upstream ...
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How to obtain the extended unaligned bases of the read in reference based genome assembly?

I did illumina read mapping based on a particular gene sequence (reference sequence) using the following command, ...
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Determining the quality of a publicly available genome assembly

I am looking for homologous genes in various protozoan species using BLAST. The genome sequences of these species are deposited in NCBI's WGS database. NCBI’s webpage includes the global statistics ...
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Coverage required

I was came across a problem during an exercise in a book and I don't really know how to solve it. I feel like something's missing. "coverage, c = $NL/G$ (N=number of reads, L=read length, G=genome ...
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Authoritative source on human cytogenetic regions?

I am looking for a database that would keep track of human cytogenetic regions and genomic coordinates per genome assembly. I had expected the Genome Reference Consortium to have it, or Ensembl, or ...
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1answer
168 views

Consensus sequence from SAM or BAM file?

I am trying to perform reference-based assembly. Most of the tutorials teach how to create a bam file and view alignemnts in IGV or Tablet. But, I want a assembled genome sequence in fasta format. How ...
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Is there a way to assemble contigs starting from a specific sequence?

My work involves searching for marker genes/fragments in metagenomic databases (like the Sequence Read Archive). Once I find these sequences, I would like to know more about the neighboring genomic ...
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Viral genome finishing

I have assembled poxvirus genome using Ray. The assembly is good. Out of several thousand contigs I got, I was able to get one scaffold using Contiguator tool, which is about 90% of my genome. I have ...
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Viral genome assembly using broad viral ngs pipeline?

I am trying to assemble RNA virus genome using Broad Viral NGS pipeline BROAD VIRAL NGS PIPELINE. I am two questions : 1) As this pipeline requires unaligned bam format as input, how do I convert ...
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Is there a resource where I can donwload E. coli assemblies grouped by their pathotypes?

I will preface this by saying I'm not a microbiologist, so I apologise for any haziness and I'll be happy to expand. I would like to download the assemblies of different E. coli genomes and group ...
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Assembly by stringtie

I run this cmd ./stringtie G1_sorted.bam -B -o G1.gtf -G Triticum_aestivum.IWGSC.42.gtf -p 4 -C G1.refs.gtf -A G1.abund.tab Error is: ...
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Is nanopolish worth it since faster polishing software is available?

For Oxford Nanopore contigs produced by any long-read assembler has anyone performed any benchmarks to compare the polishing tools racon, ...
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Separation of mixed plasmid DNA sequences post whole-plasmid sequencing

Imagine a DNA sample containing a mixture of different intact plasmids. These samples are sequenced using either MiSeq or HiSeq sequencing. Would it possible to assemble these plasmids post-sequencing ...
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How to get fasta alignment file from SAM/BAM file?

I am not talking about consensus sequence, I know how to get consensus sequence using mpileup in samtools/bcftools. As I understand , SAM/BAM files are basically sequence alignment format so it's ...
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wtdbg2: practical implications of k-mer fsize and psize choice

I am using wtdbg2 2.3 to assemble a human genome (sequenced on PromethION from a cell line). I filtered out reads with low average quality, and now I am trying to determine the parameters that will ...
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How is consensus alignment for OLC assembly usually implemented?

Background In C++ I want to implement a simple class that performs overlap layout consensus assembly but I can not figure out the most logical data structure to use for the consensus alignment step. ...
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Calculating alignment/mapping time

I am trying to assemble a plant genome using AWS resources using velvet. Plant genome is huge (> 10 times human genome) and coverage is around 30 x. We are planning for de novo assembly with Velvet (...
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1answer
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E.coli Sequencing & Analysis

I have been given the task of assembling a 'new' Ecoli genome and analysing the genes present etc. The Ecoli is a new strain, and has been taken and run on a Nextseq 500 in high-output mode with ...
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Problem with SPAdes installation - no bin folder

I have been trying to download SPAdes for an important project I wanted to do, however, I have encountered a small problem while doing so. It appears that there is no 'bin' folder in my SPAdes ...
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How many reads do I need for hybrid assembly

I have Illumina and PacBio reads and I would like to use dbg2olc for hybrid assembly. Part of dbg2olc is SelectLongestReads which select reads that sum to ...
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How to assess the quality of assembled .fasta genome files?

I have assembled 3 .fasta files from contigs infastq format of 3 different Homo sapiens. I would like to see if the assembled ...
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estimate genome size: kmer-based approach from PacBio reads

Can anyone suggest a software/method for kmer analysis using PacBio reads (RSII)? Something similar to Jellyfish, that I saw in a nice tutorial - but must be suitable for long, noisy reads. ...
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What is the current state-of-the-art in assembling hybrid transcriptomes?

We are considering attempting de novo assembly of a species transcriptomes (i.e. without a reference genome) using the combined NGS outputs of Iso-seq and Illumina. One example I saw (Li et al 2017),...
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Merge / Reconciliate several de novo transcriptome assemblies with different kmers

I am trying to build a De Novo transcriptome reference assembly for an eukaryotic organism. I've created several assemblies with rnaSpades using different kmer sizes (19 to 69 with step 10). Now I ...
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Find paralogs in a draft genome

We generated a (diploid, chordata, highly heterozgous) genome using PacBio and we wanted to see whether it contains lineage-specific duplications (paralogs, basically). The genome is not in Ensembl ...
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Reduce number of transcripts in a highly variable de novo transcriptome assembly

I have a de novo assembly using both multiple SRA and locally sequenced transcriptomes. I started with 270M PE reads from 9 tissues. Here are the assembly stats generated with ...
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Correct way to assemble reference transcriptome - what's --samples_file option in Trinity?

I have 101 samples from 9 tissues of a nonmodel species, coming either from SRA or sequenced by my lab. I want to generate a reference transcriptome with Trinity for further analysis. In order to do ...
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1answer
104 views

Viral Metagenomics

I am analyzing viral metagenomics data (Illumina Miseq) for the first time. I have used Ray for de novo viral genome assembly before but I haven't done metagenomics analysis before. I know that ...
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Why isn't there a standard method to convert GFA to JSON?

I'm confused why the GFA format doesn't have a standard JSON parser. It appears to be related to a decision made several years ago, here. The objection appears to be "JSON isn't necessary, as it's ...
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Is there a standard definition for “assembly polishing”?

Is there a standard definition for "assembly polishing" in the field? Is there a standard definition for what polishing algorithms do? My understanding of "polishing" is strongly influenced by ...
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Does the galGal5 chicken assembly have a chromosome 29?

The chromosome sizes at UCSC don't seem to contain chr29: ftp://hgdownload.soe.ucsc.edu/goldenPath/galGal5/bigZips/galGal5.chrom.sizes It has a chr28 and a chr30. Am I missing something or is there ...
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283 views

Cufflinks Error: sort order of reads in BAMs must be the same

I am running Cufflinks for transcriptome assembly using the .bam file generated by Hisat2. I tried both bam and sorted bam files ...
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Canu failed with 'didn't find any mers?'

I have installed canu using conda in an environment containing Python 3 on a Mac OS X. I checked that canu is properly installed by pulling out the help and it works. I launched a canu assembly job by ...
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Why can using more reads lead to a lower quality assembly?

I am experimenting with adding additional reads to the input files I'm giving SOAPdenovo2, and there comes a point where a good contig I've been watching actually stops showing up. Does anyone have a ...
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Make ipyrad use cuda-enabled NVidia card on Ubuntu

I want to use ipyrad on a new Ubuntu machine that has an NVidia Quadro K2000 card with 384 cores. One can configure ipyrad to run on a linux cluster. Do I have any options to get ipyrad to access ...
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Difference between de novo transcriptome assembly methods

I have been looking around (including read the original papers) to understand what is essentially the difference between StringTie in non-reference based mode (de novo) and Trinity de novo assembly. I ...
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Can the canu assembler output a fastq file of the final assembly just like HGAP4?

I have assembled some genome from Sequel PacBio data both with HGAP4 on the SMRT Link interface and using canu on the command line. The HGAP4 assembler outputs a fastq file of the final assembly such ...
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SPAdes error during assembly

I try to perform a hybrid assembly using Unicycler. Unicycler use SPAdes to assembly the illumina sequences, and the program crashes while in SPAdes. The spades.log file says something about an ...
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1answer
106 views

Scaffolding a genome with hybrid data

I am assembling a ~500MB genome, and have ~150x long reads and ~200x 150bp PE short reads, with a ~400bp insert size. I've done a lot of work with minimap+miniasm, and have what I think is a good set ...
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How to submit a canu job on LSF high-performance computing cluster farm?

I am currently running canu on an LSF Linux server using the following script called assemble.sh: ...
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1answer
194 views

Java error when launching Pilon

I am trying to use Pilon to improve a reference based on some Illumina data I have got. So, I aligned the Illumina reads to the reference using bwa. Then, I want to use Pilon to improve the reference ...
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142 views

Verify a predicted protein in one genome in a different genome of the same species

I have two genome assemblies of the same non-model species, call them Assembly 1 (generated from Illumina data) and Assembly 2 (generated from PacBio data). For Assembly 1, I also have predicted ...
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Genome assembly from error-prone reads

I understand how to assemble genome from error-free reads. I implemented like this: Construct directed overlap graph with reads as vertices and edges as maximum overlap between two vertices. ...
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How to calculate overall reference coverage with MUMmer?

Is the MUMmer suite capable of calculating reference sequence coverage statistics for all query sequences collectively? It would be possible to achieve by parsing the output of ...
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Reordering scaffolds according to a reference without a genetic map

I am trying to reorder scaffolds of a rice species, but no genetic map is available right now. Oryza sativa Japonica is a close relative of this rice species. Mummer was used to do a whole genome ...