BOL: Related items

Understanding PacBio

Jitendra Narayan — Fri, 24 Feb 2017 10:17:36 -0600

This tutorial includes resources for learning more about PacBio data and bioinformatics analysis, and includes content suitable for both beginners and experts. Below are links to training modules (webinars and PowerPoint presentations) to help you get started with your data processing, as well as information for specialized applications.

Training Resources:

Specialized Applications:

Address of the bookmark: https://github.com/PacificBiosciences/Bioinformatics-Training/wiki

Ribbon !!

Jit — Fri, 21 Oct 2016 04:54:30 -0500

Visualization has played an extremely important role in the current genomic revolution to inspect and understand variants, expression patterns, evolutionary changes, and a number of other relationships. However, most of the information in read-to-reference or genome-genome alignments is lost for structural variations in the one-dimensional views of most genome browsers showing only reference coordinates. Instead, structural variations captured by long reads or assembled contigs often need more context to understand, including alignments and other genomic information from multiple chromosomes. We have addressed this problem by creating Ribbon (genomeribbon.com) an interactive online visualization tool that displays alignments along both reference and query sequences, along with any associated variant calls in the sample. This way Ribbon shows patterns in alignments of many reads across multiple chromosomes, while allowing detailed inspection of individual reads (Supplementary Note 1). For example, here we show a gene fusion in the SK-BR-3 breast cancer cell line linking the genes CYTH1 and EIF3H. While it has been found in the transcriptome previously, genome sequencing did not identify a direct chromosomal fusion between these two genes. After SMRT sequencing, Ribbon shows that there are indeed long reads that span from one gene to the other, going through not one but two variants, for the first time showing the genomic link between these two genes (Figure 1a). More gene fusions of this cancer cell line are investigated in Supplementary Note 2. Figure 1b shows another complex event in this sample made simple in Ribbon: the translocation of a 4.4 kb sequence deleted from chr19 and inserted into chr16 (Figure 1b). Thus, Ribbon enables understanding of complex variants, and it may also help in the detection of sequencing and sample preparation issues, testing of aligners and variant-callers, and rapid curation of structural variant candidates (Supplementary Note 3). In addition to SAM and BAM files with long, short, or paired-end reads, Ribbon can also load coordinate files from whole genome aligners such as MUMmer. Therefore, Ribbon can be used to test assembly algorithms or inspect the similarity between species. Supplementary Note 4 shows a comparison of gorilla and human genomes using Ribbon, highlighting major structural differences. In conclusion, Ribbon is a powerful interactive web tool for viewing complex genomic alignments.

Script at https://github.com/MariaNattestad/ribbon

Address of the bookmark: http://genomeribbon.com/

LINKS

Jit — Fri, 04 Nov 2016 06:19:01 -0500

LINKS is a genomics application for scaffolding genome assemblies with long reads, such as those produced by Oxford Nanopore Technologies Ltd. It can be used to scaffold high-quality draft genome assemblies with any long sequences (eg. ONT reads, PacBio reads, another draft genomes, etc)

Paper at https://gigascience.biomedcentral.com/articles/10.1186/s13742-015-0076-3

Address of the bookmark: https://github.com/warrenlr/LINKS/

SGA: String Graph Assembler

Jit — Thu, 08 Dec 2016 05:08:59 -0600

SGA is a de novo genome assembler based on the concept of string graphs. The major goal of SGA is to be very memory efficient, which is achieved by using a compressed representation of DNA sequence reads.

More at

https://github.com/jts/sga

SGA dependencies:
-google sparse hash library (http://code.google.com/p/google-sparsehash/)
-the bamtools library (https://github.com/pezmaster31/bamtools)
-zlib (http://www.zlib.net/)
-(optional but suggested) the jemalloc memory allocator (http://www.canonware.com/jemalloc/download.html)

Address of the bookmark: https://github.com/jts/sga

Structural variation: the hidden genomic treasure

Jit — Sat, 10 Dec 2016 16:19:09 -0600

Genome re-sequencing projects have revealed substantial amounts of genetic variation between individuals extending beyond single nucleotide polymorphisms (SNPs) and short indels. Structural Variations (SVs) and Copy Number Variations (CNVs) are a major source of genomic variation. However, compared to SNPs, accurate detection, genotyping and understanding of CNVs is lagging behind due to much greater analytical challenges related to SV/CNV detection and analysis. In our lab we analyse SVs/CNVs using high-throughput sequencing and different analytical approaches. The most‐studied structural variants are copy number variations (CNVs) which can be generated by several different mechanisms including non‐allelic homologous recombination, non‐homologous end‐joining and deoxyribonucleic acid (DNA) replication‐related fork stalling and template switching. CNVs are closely related to segmental duplications (SDs): SDs can stimulate the formation of CNVs and themselves started out as CNVs, but became fixed in a species. Structural variation can be neutral but has also influenced our phenotypic evolution, for example our susceptibility to disease and our ability to digest certain types of food. Our understanding of the extent of structural variation is increasing rapidly, but it will be much more difficult to understand its phenotypic consequences.

Structural variants (SVs) such as deletions, insertions, duplications, inversions and translocations litter genomes and are often associated with gene expression changes and severe phenotypes (ie. genetic diseases in humans). Recent studies on the functional aspects of different types of SVs have unveiled several cases of adaptive evolution. For example, inversions have been associated with ecological adaptations and may facilitate speciation. Due to their prevalent nature, SVs arguably have a large impact on genome evolution and should not be neglected when studying the genetics of adaptation and speciation. SVs were classically defined as chromosomal rearrangements larger than 1kb, but due to a higher resolution of new detection methods, smaller variants (between 50 and 1000 base pairs) can now be accurately assessed. Besides various methods of detection in next generation sequencing data (paired end mapping, split reads, and depth of coverage), array-based approaches have proven to be particularly useful for detecting copy number variations (CNVs). These technologies have enabled researchers to catalog a wide spectrum of SVs in many organisms and infer the effects of selection shaping their evolutionary trajectories.

Structure variation sequencing signature (Source: NatRev Genetics)

Related tools, databases and publications are listed below. If you know any interesing papers, please let us know in comment section:

Key concepts

Structural variation includes balanced variants such as inversions and translocations, and unbalanced ones such as duplications and deletions (copy number variations or CNVs).

Structural variants can arise by several mechanisms, including nonallelic homologous recombination (NAHR), nonhomologous end‐joining (NHEJ) and DNA replication‐based fork stalling and template switching (FoSTeS).

CNV is closely linked to segmental duplication, but is not exactly the same. Segmental duplications can stimulate CNV formation by NAHR, and themselves arise from CNVs that have become fixed.

Segmental duplications did not appear uniformly during the evolution of the Great Ape species, but rather during a burst of activity around the time of the divergence of gorilla from the human/chimpanzee ancestor.

Duplicated genes play a critical role in the evolution of a genome as they act as ‘spare parts’ than can evolve to perform new or more specialized functions.

Effects of structural variation on gene expression can be identified but only a few examples of the consequences for species biology have been documented.

Tools

CNVnatora tool for CNV discovery and genotyping from depth of read mapping.2011a,2011b

AGEa tools that implements an algorithm for optimal alignment of sequences with SVs.2011

BreakSeqa pipeline for annotation, classification and analysis of SVs at single nucleotide resolution.2010

PEMera computational and simulation framework for discovering SVs by paired-end read mapping.2009,2007

GASV https://code.google.com/archive/p/gasv/

PAIROSCOPE http://pairoscope.sourceforge.net/

SVDetect http://svdetect.sourceforge.net/Site/Home.html

BreakPtr, discovery of unbalanced structural variants (copy-number variants) with tiling microarrays Link

R Package https://www.bioconductor.org/help/course-materials/2010/EMBL2010/Practical-4-StructuralVariants.pdf

BreakSeq, structural variant genotyping using split reads Link

CopySeq, genotyping of unbalanced structural variants (copy-number variants) using read-depth Link

DELLY2, integrated structural variant discovery, genotyping and visualization in deep sequencing data Link

PEMer, structural variant discovery in 454 sequencing data by paired-end mapping Link

TIGER, transduction inference in germline genomes using short read data Link

MANTA https://github.com/Illumina/manta

SV-Bay https://github.com/InstitutCurie/SV-Bay

BreakDancer http://breakdancer.sourceforge.net/

Variation Hunter http://compbio.cs.sfu.ca/software-variation-hunter

Lumpy https://github.com/arq5x/lumpy-sv

ForestSV http://sebatlab.ucsd.edu/index.php/software-data

PBSuites for long reads https://sourceforge.net/projects/pb-jelly/

Visualization

The SV visualization tool: http://genomesavant.com/savant/

InGAP-SV (http://ingap.sourceforge.net/) that is nice tools for both detection and visualisation of severals kind of structural variations (Large insertions, translocation, deletion, inversions....)

Tools table: http://www.nature.com/nbt/journal/v29/n8/fig_tab/nbt.1904_T2.html

Variation Viewer https://www.ncbi.nlm.nih.gov/variation/view/

Papers

http://www.nature.com/nmeth/journal/v9/n2/full/nmeth.1858.html

http://journal.frontiersin.org/researchtopic/1412/structural-variations-in-genomes-ecological-and-evolutionary-implications

http://www.mi.fu-berlin.de/wiki/pub/ABI/GenomicsLecture10Materials/structural-variation.pdf

http://bmcgenomics.biomedcentral.com/articles/10.1186/s12864-015-1479-3

https://www.ncbi.nlm.nih.gov/dbvar/content/overview/

http://www.nature.com/subjects/structural-variation

https://eichlerlab.gs.washington.edu/news/NatMeth_Feb2012.pdf

https://www.ncbi.nlm.nih.gov/pubmed/19477992 ***

https://www.ncbi.nlm.nih.gov/pubmed/22452995

http://biorxiv.org/content/early/2016/09/06/073833

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4479793/

http://www.nature.com/articles/srep18501

http://www.genetics.org/content/202/1/351

http://www.cs.cmu.edu/~sssykim/teaching/s13/slides/Lecture_SVI.pdf

https://www.omicsonline.org/open-access/structural-variation-detection-from-next-generation-sequencing-2469-9853-S1-007.php?aid=69055

http://schatzlab.cshl.edu/presentations/2016/2016.01.12.PAG.Structural%20Variations.pdf

CISA: Contig Integrator for Sequence Assembly

Neel — Thu, 15 Dec 2016 05:42:21 -0600

A plethora of algorithmic assemblers have been proposed for the de novo assembly of genomes, however, no individual assembler guarantees the optimal assembly for diverse species. Optimizing various parameters in an assembler is often performed in order to generate the most optimal assembly. However, few efforts have been pursued to take advantage of multiple assemblies to yield an assembly of high accuracy. In this study, we employ various state-of-the-art assemblers to generate different sets of contigs for bacterial genomes. A tool, named CISA, has been developed to integrate the assemblies into a hybrid set of contigs, resulting in assemblies of superior contiguity and accuracy, compared with the assemblies generated by the state-of-the-art assemblers and the hybrid assemblies merged by existing tools. This tool is implemented in Python and requires MUMmer and BLAST+ to be installed on the local machine. The source code of CISA and examples of its use are available at http://sb.nhri.org.tw/CISA/.

Address of the bookmark: http://sb.nhri.org.tw/CISA/en/CISA

GARM:Genome Assembly, Reconciliation and Merging

Jit — Mon, 19 Dec 2016 06:03:02 -0600

The pipeline is based mainly implemented using Perl scripts and modules and third-party open source software like the AMOS (Myers et al., 2000) and MUMmer (Kurtz et al., 2004) packages. The pipeline was tested on Debian, Ubuntu, Fedora and BioLinux distributions. The method merges contigs or scaffolds from different assemblers using the same or different sequencing technologies. When scaffolds are provided, a process of finding probable compressions or extensions (CE) problems in the assemblies can be per-formed; contigs are joined back into scaffolds after gap recalculation

Address of the bookmark: http://garm-meta-assem.sourceforge.net/

quickmerge: A simple and fast metassembler and assembly gap filler designed for long molecule based assemblies.

Jit — Mon, 19 Dec 2016 10:23:36 -0600

quickmerge uses a simple concept to improve contiguity of genome assemblies based on long molecule sequences, often with dramatic outcomes. The program uses information from assemblies made with illumina short reads and PacBio long reads to improve contiguities of an assembly generated with PacBio long reads alone. This is counterintuitive because illumina short reads are not typically considered to cover genomic regions which PacBio long reads cannot. Although we have not evaluated this program for assemblies generated with Oxford nanopore sequences, the program should work with ONP-assemblies too.

Address of the bookmark: https://github.com/mahulchak/quickmerge

Genome Assembly Tutorial

Abhimanyu Singh — Tue, 20 Dec 2016 07:56:01 -0600

If genomes were completely random sequences in a statistical sense, 'overlap-consensus-layout' method would have been enough to assemble large genomes from Sanger reads. In contrast, real genomes often have long repetitive regions, and they are hard to assemble using overlap-consensus-layout approach. De Bruijn graph-based assembly approach was originally proposed to handle the assembly of repetitive regions better.

More at http://www.homolog.us/Tutorials/index.php?p=1.4&s=1

Address of the bookmark: http://www.homolog.us/Tutorials/index.php?p=1.4&s=1

Progressive Cactus

Jit — Tue, 17 Jan 2017 03:40:06 -0600

v0.0 by Glenn Hickey (hickey@soe.ucsc.edu)

Progressive Cactus is a whole-genome alignment package.

Requirements

git
gcc 4.2 or newer
python 2.7
wget
64bit processor and build environment
150GB+ of memory on at least one machine when aligning mammal-sized genomes; less memory is needed for smaller genomes.
Parasol or SGE for cluster support.
750M disk space

Instructions

IMPORTANT NOTE: Progressive Cactus does not presently support installation into paths that contain spaces. Until this is resolved, you can use a softlink as a workaround: ln -s "path with spaces" "installation path without spaces"

In the parent directory of where you want Progressive Cactus installed:

git clone git://github.com/glennhickey/progressiveCactus.git
cd progressiveCactus
git pull
git submodule update --init
make

It is also convenient to add the location of progressiveCactus/bin to your PATH environment variable. In order to run the included tools (ex hal2maf) in the submodules/ directory structure, first source progressiveCactus/environment to load the installed environment.

If any errors occur during the build process, you are unlikely to be able to use the tool. Please submit a GitHub issue so we can help out: not only will you help yourself, but others who wish to use the tool as well.

Note that all dependencies are also built and included in the submodules/ directory. This increases the size and build time but greatly simplifies installation and version management. The installation does not create or modify any files outside the progressiveCactus/ directory.

Address of the bookmark: https://github.com/glennhickey/progressiveCactus