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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Difference between genome assembly and genome sequence alignment to a reference to find structural variants

I'm trying to determine what the difference and benefits of genome assembly and genome sequence alignments are when trying to identify structural variants or transposons in populations. I've been ...
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How to fragment genomes into non-overlapping sequences of differing sizes?

I have bacterial chromosomes/plasmids that I have downloaded off refseq in a multi-fasta that I would like to fragment into non-overlapping fragments of between 1 kb and 15 kb. I have tried googling ...
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Bacterial DNA at the tail of transcriptome reads. What does that mean?

I am assembling a transcriptome obtained from the Internet. The transcriptome was extracted from a human cancer tissue that had been previously grafted into a mouse. I have detected that many ...
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Comparing gene abundances between metagenomes

My workflow until now: Find fragments of a marker gene in unassembled metagenomes > download and assemble metagenomes > recover the gene neighborhood / gene set of interest Right now I have a ...
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Can SBOL encode DNA Assembly linkers?

Where in an SBOL file could you specify the linkers that would be placed as prefixes / suffixes between parts for assembly? Lets say you have a final construct of your design and want to assemble the ...
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60 views

Decontaminating RNA seqs for de novo transcriptome assembly and annotation of novel eukaryotes

I have raw paired-end RNA-seq reads for two novel eukaryotic species. Some background: the reads represent a copepod (arthropod) species each. The mRNA for each read set was obtained by extracting ...
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Gffcompare issue: 0 reference transcripts loaded

I am trying to use gffcompare to compare my assembled transcriptome to a reference gtf that contains information about small open reading frames (sORFs). The reference gtf was obtained by processing ...
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1answer
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“megahit: command not found” after correct installation with miniconda

I want to use MEGAHIT (git: https://github.com/voutcn/megahit) to assemble metagenome shotgun sequencing reads (illumina). I install the package with the command <...
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Methods to predict one connected structure from two known separate structures

Currently, I am facing such a question: how to predict one connected protein structures from two separate known structures? It should be different from the protein-protein docking method, which is ...
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Assemble reads for a specific gene

I have a lot of unassembled sequencing data and I want to build phylogeny for some genes from these data. I can retrieved the sequene by mapping reads to a reference sequence but if the two species ...
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After KEGG and GO analysis, how to make tables+phylogenetic trees

hope everyone is ok. I used Trinity to do a de novo transcriptome assembly, then blastp/blastx and then used Blast2GO software to do KEGG and GO analysis. So i got some txt files with header : for GO ...
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What is the meaning of these misaligned reads in a sequencing run?

I guess that's a basic question but I can't find an answer. I am analyzing some SARS-CoV-2 sequencing runs. I often find read alignments like the one in the image. ...
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Platanus-allee phasing fail: Error(13): Error, SolveDBG exception!

I am using Platanus-allee 2.2.2 for heterozygous genome (~500mb) assembly with Illumina short reads and PacBio reads input data. I have the file contigs.fa from short reads but phasing step with ...
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Is there any value in scaffolding the output contigs of MEGAHIT assembler given a metagenomic dataset?

As far as I understood, for most assembly programs, the scaffolding step takes into consideration paired-end information in order to get from contigs (contiguous sequences) to scaffolds (longer ...
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Will using smaller kmers help get larger contigs? If not, then what?

I've been using MEGAHIT to assemble metagenomes, with particular focus on specific genomic areas. Sometimes all I get is gene fragments or pathway fragments (eg. if I know that genes A, B, C, D and E ...
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polishing assembled genome to QV50 value

I am trying to assemble some genomes from pacbio reads using HGAP3 followed by quiver polishing. However, even after multiple rounds of quiver polishing, I cannot attain quality value 50 (QV50) for a ...
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Optimum parameters for genome assembly via canu

I have been trying to assemble my genome via canu. Canu gave me about 200 contigs using default parameters. My genomesize is ~9Mbp. Can you recommend some parameters I can tweak to decrease the ...
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Tools for comparing/visualizing FASTAs?

I have two FASTAs/assemblies of the same species, but the bases are somewhat different. I would like to explore this. What tools/methods exist to compare two FASTAs and see the difference in ...
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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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118 views

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
223 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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1answer
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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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129 views

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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2answers
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Merge / Reconciliate several de novo transcriptome assemblies with different kmers

I am building a De Novo transcriptome reference assembly for an eukaryotic organism for which I have a genome. I've created several assemblies with rnaSpades using different kmer sizes (19 to 69 with ...
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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 ...