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Questions tagged [pacbio]

Pacific Biosciences is a biotech company selling a third-generation sequencing technology often denoted by the same name, also known as Single-Molecule Real-Time (SMRT) sequencing. Questions related to data generated by PacBio/SMRT sequencing.

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43 views

Best way to “fish” long reads that match a query sequence

Very simple set-up: We have a PacBio long high-fidelity (HiFi) reads, genome sequencing, and some of those that contain a particular sequence. I want to find out which ones. The length of the query ...
5
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1answer
90 views

Why is a PacBio read length larger than the aligned reference region?

I recently had some Iso-Seq sequencing done on my organism catfish on the new Sequel platform and got weird alignments for a size selected 4 Kilobase and up fraction after running the isoseq3 pipeline....
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2answers
175 views

Mapping Reads to Known Gene Paralogs with Long Read Technology

I have some sequencing data from a captured region that is a known paralog edited. For now, I have been mapping the data using standard minimap2 flags for PacBio DNA sequencing: ...
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2answers
169 views

calling diploid SNVs from long reads

I'd like to call diploid SNV variants from long-reads data (~80SMRTcells PacBio). I have generated a draft reference genome for an indivudual from a heterozygous (~4%) species (Canu+Haplomerger2). ...
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1answer
49 views

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 ...
8
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1answer
602 views

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. ...
4
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0answers
43 views

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),...
2
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1answer
443 views

Can I pilon-polish long reads with Illumina short reads to improve structural variant detection?

I have pacbio Sequel data at 50 x coverage for a strain of animal. I would like to find structural variants compared to the reference genome sequence. At the moment, I align my reads to the reference ...
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0answers
43 views

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 ...
5
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1answer
81 views

Can pair end reads with high heterozygosity be used to polish PacBio assembly?

I have used canu (correct) and smartdenovo to assembly PacBio reads. Next I am going to polish my assembly using illumina pair end reads by Pilon. However, I found there is high heterozygosity in my ...
2
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1answer
112 views

How to interpret the SMRT Link base modification algorithm output?

I have got PacBio data from a Sequel machine at a coverage > 50X. I have run it in SMRT Link with their Base Modification Detection algorithm. I am now trying to figure out how to interpret the output ...
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3answers
208 views

How to install blasr on Mac OS X?

I am trying to install blasr on mac osx. I followed the steps here I get the error below after using the 'make' command: ...
2
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1answer
106 views

Is it possible to find out the PacBio chemistry from the PacBio Sequel BAM's header?

Is it possible to find out the PacBio chemistry from the PacBio Sequel BAM's header? E.g. whether it was produced with P5-C3 or P6-C4? I checked mine but I haven't found it but maybe it's hidden in ...
7
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1answer
445 views

Coverage calculation: long reads (RNA-seq)

Say your aim is to calculate the coverage of an RNA-seq experiment generated with long-read sequencing (so, uneven read length). Up to now, I relied on the Lander/Waterman equation: $$C = L*N / G$$...
9
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1answer
1k views

Which quality score encoding does PacBio use?

Do you know which quality score encoding PacBio uses now? I know some of their file formats have changed in the past year or two, but I haven't found much on their quality score encoding. The most ...
6
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2answers
137 views

Structural variant calling for low-coverage PacBio data

PacBio is selling ~10x PacBio SEQUEL long reads as an upgrade to Illumina data for SV discovery. In a clinical setting, the main requirements are proper sensitivity and specificity but also the ...