BOL: Related items

Prokka: tool for the rapid annotation of prokaryotic genomes

Jit — Mon, 06 Mar 2017 03:49:57 -0600

Prokka is a software tool for the rapid annotation of prokaryotic genomes. A typical 4 Mbp genome can be fully annotated in less than 10 minutes on a quad-core computer, and scales well to 32 core SMP systems. It produces GFF3, GBK and SQN files that are ready for editing in Sequin and ultimately submitted to Genbank/DDJB/ENA.

Address of the bookmark: http://www.vicbioinformatics.com/software.prokka.shtml

COCACOLA (binning metagenomic contigs using sequence COmposition, read CoverAge, CO-alignment, and paired-end read LinkAge)

Jit — Tue, 07 Mar 2017 08:50:57 -0600

COCACOLA is a general framework that combines different types of information: sequence COmposition, CoverAge across multiple samples, CO-alignment to reference genomes and paired-end reads LinkAge to automatically bin contigs into OTUs. Furthermore, COCACOLA seamlessly embraces customized prior knowledge to facilitate binning accuracy.

News: Python version of COCACOLA is available now!

Address of the bookmark: https://github.com/younglululu/COCACOLA

Multigenome assembly

Jit — Tue, 14 Mar 2017 04:41:23 -0500

This project contains scripts and tutorials on how to assemble individual microbial genomes from metagenomes, as described in:

Genome sequences of rare, uncultured bacteria obtained by differential coverage binning of multiple metagenomes

Mads Albertsen, Philip Hugenholtz, Adam Skarshewski, Gene W. Tyson, Kåre L. Nielsen and Per .H. Nielsen

Nature Biotechnology 2013, doi: 10.1038/nbt.2579

See the associated online guide for detailed information.

https://github.com/MadsAlbertsen/multi-metagenome

Address of the bookmark: https://github.com/MadsAlbertsen/multi-metagenome

Download assemblies from NCBI

Bulbul — Mon, 15 May 2017 06:02:32 -0500

A new “Download assemblies” button is now available in the Assembly database. This makes it easy to download data for multiple genomes without having to write scripts.

For example, you can run a search in Assembly and use check boxes (see left side of screenshot below) to refine the set of genome assemblies of interest. Then, just open the “Download assemblies” menu, choose the source database (GenBank or RefSeq), choose the file type, and start the download. An archive file will be saved to your computer that can be expanded into a folder containing your selected genome data files.

More at https://ncbiinsights.ncbi.nlm.nih.gov/2017/05/08/genome-data-download-made-easy/

CAMSA :: a tool for Comparative Analysis and Merging of Scaffold Assemblies

Rahul Nayak — Thu, 28 Dec 2017 09:10:26 -0600

CAMSA – is a tool for Comparative Analysis and Merging of Scaffold Assemblies, distributed both as a standalone software package and as Python library under the MIT license.

Main features:

works with any number of scaffold assemblies in de-novo non-progressive fashion
allows to simultaneously work with scaffold assemblies obtained from any in silico and in vitro techniques, supporting multiple existing formats via built-in converters
creates an extensive report with several comparative quality metrics (both on assembly level and on the level of individual assembly points)
constructs a merged combined scaffold assembly
provides an interactive framework for a visual comparative analysis of the given assemblies

Address of the bookmark: https://cblab.org/camsa/

RNA-seq Analysis Workshop Course Materials

Jit — Tue, 03 Jul 2018 08:14:14 -0500

RNAseq can be roughly divided into two "types": Reference genome-based - an assembled genome exists for a species for which an RNAseq experiment is performed. It allows reads to be aligned against the reference genome and significantly improves our ability to reconstruct transcripts. This category would obviously include humans and most model organisms but excludes the majority of truly biologically intereting species (e.g., Hyacinth macaw); Reference genome-free - no genome assembly for the species of interest is available. In this case one would need to assemble the reads into transcripts using de novo approaches. This type of RNAseq is as much of an art as well as science because assembly is heavily parameter-dependent and difficult to do well. In this lesson we will focus on the Reference genome-based type of RNA seq. http://chagall.med.cornell.edu/RNASEQcourse/

Address of the bookmark: http://chagall.med.cornell.edu/RNASEQcourse/

GWASpro: A High-Performance Genome-Wide Association Analysis Server

Rahul Nayak — Fri, 07 Dec 2018 08:04:57 -0600

GWASpro supports building complex design matrices, by which complex experimental designs that may include replications, treatments, locations and times, can be accounted for in the linear mixed model (LMM). GWASpro is optimized to handle GWAS data that may consist of up to 10 million markers and 10,000 samples from replicable lines or hybrids. GWASpro provides an interface that significantly reduces the learning curve for new GWAS investigators.

Address of the bookmark: https://bioinfo.noble.org/GWASPRO/

Sequence Ontology Bioinformatics Analysis (SOBA) tool to provide a simple statistical and graphical summary of an annotated genome

BioStar — Wed, 22 Jul 2020 10:11:13 -0500

We have developed the Sequence Ontology Bioinformatics Analysis (SOBA) tool to provide a simple statistical and graphical summary of an annotated genome. We envisage its use during annotation jamborees, genome comparison and for use by developers for rapid feedback during annotation software development and testing. SOBA also provides annotation consistency feedback to ensure correct use of terminology within annotations, and guides users to add new terms to the Sequence Ontology when required. SOBA is available at http://www.sequenceontology.org/cgi-bin/soba.cgi.

More at https://pubmed.ncbi.nlm.nih.gov/20494974/

Address of the bookmark: http://www.sequenceontology.org/cgi-bin/soba.cgi

Bactopia: a Flexible Pipeline for Complete Analysis of Bacterial Genomes

Abhi — Wed, 15 May 2024 14:36:12 -0500

Bactopia is a flexible pipeline for complete analysis of bacterial genomes. The goal of Bactopia is to process your data with a broad set of tools, so that you can get to the fun part of analyses quicker!

Bactopia can be split into two main parts: Bactopia Analysis Pipeline, and Bactopia Tools.

Bactopia Analysis Pipeline is the main per-isolate workflow in Bactopia. Built with Nextflow, input FASTQs (local or available from SRA/ENA) are put through numerous analyses including: quality control, assembly, annotation, minmer sketch queries, sequence typing, and more.

Bactopia Tools are a set a independent workflows fo

Address of the bookmark: https://github.com/bactopia/bactopia

RNA-Seq Analysis: A Guide for Bioinformaticians

LEGE — Sat, 07 Dec 2024 22:22:24 -0600

RNA sequencing (RNA-Seq) has revolutionized transcriptomics, offering unprecedented insights into gene expression, splicing, and transcript diversity. For bioinformaticians, RNA-Seq analysis is a gateway to exploring the complexity of RNA biology and its implications in health and disease. This blog post provides an overview of RNA-Seq analysis, key computational steps, and tools for bioinformaticians eager to delve into this powerful technique.

What is RNA-Seq?

RNA-Seq is a next-generation sequencing (NGS) technology used to study the transcriptome—the complete set of RNA molecules in a cell. It quantifies gene expression, detects novel transcripts, and captures alternative splicing events with high sensitivity and resolution.

Workflow for RNA-Seq Analysis

RNA-Seq analysis involves several stages, each requiring computational tools and expertise.

1. Experimental Design and Data Acquisition

Before diving into analysis, bioinformaticians should consider:

Biological Replicates: Ensure statistical power to detect meaningful differences.
Sequencing Depth: Align sequencing depth to study objectives (e.g., higher depth for low-abundance transcripts).
Paired-End vs. Single-End: Paired-end sequencing provides more detailed information on transcript structure.

Once sequencing is complete, raw data is provided in FASTQ format, containing sequence reads and quality scores.

2. Quality Control and Preprocessing

Quality control (QC) ensures data integrity. Tools such as FastQC evaluate metrics like base quality, GC content, and adapter contamination.

Preprocessing Steps:

Trimming: Tools like Trimmomatic or Cutadapt remove low-quality bases and adapter sequences.
Filtering: Discard reads below a certain quality threshold or length.

3. Read Alignment

Reads are mapped to a reference genome or transcriptome to determine their origin. Alignment tools include:

HISAT2: Handles large genomes efficiently and supports spliced alignments.
STAR: High-speed aligner optimized for RNA-Seq.
Bowtie2: Suitable for short-read alignment.

Output: A SAM/BAM file containing aligned reads.

4. Transcript Assembly and Quantification

This step involves identifying transcripts and quantifying their expression levels. Tools used include:

StringTie: Assembles and quantifies transcripts from aligned reads.
Salmon/Kallisto: Perform pseudo-alignment for rapid and accurate quantification.

Expression levels are typically measured as TPM (transcripts per million) or FPKM (fragments per kilobase of transcript per million mapped reads).

5. Differential Expression Analysis

To identify genes with altered expression between conditions, bioinformaticians use tools such as:

DESeq2: Accounts for data normalization and variability.
edgeR: Handles overdispersed count data efficiently.
Limma-voom: Combines linear modeling with RNA-Seq count data.

The output includes a list of differentially expressed genes (DEGs) with statistical significance and fold-change values.

6. Functional Annotation and Pathway Analysis

Understanding the biological significance of DEGs involves:

Gene Ontology (GO) Analysis: Tools like DAVID or clusterProfiler categorize genes based on their biological functions.
Pathway Enrichment Analysis: Identifies pathways enriched in DEGs using tools like KEGG, Reactome, or GSEA.

7. Visualization

Visualizing results enhances interpretability. Common visualizations include:

Heatmaps: Show expression patterns across samples (e.g., pheatmap).
Volcano Plots: Highlight significant DEGs (e.g., ggplot2).
PCA/UMAP: Assess sample clustering and variability (e.g., Seurat).

Challenges in RNA-Seq Analysis

Batch Effects: Technical variability can confound biological signals. Combat this with normalization techniques or batch-correction tools like ComBat.
Low-Quality Samples: Poor-quality RNA impacts downstream analyses.
Computational Complexity: RNA-Seq generates massive datasets, requiring robust computing resources and optimized pipelines.

Key Tools and Resources

Bioconductor: A treasure trove of R packages for RNA-Seq analysis.
Galaxy: A web-based platform for running RNA-Seq workflows.
Nextflow/Snakemake: Workflow management tools to streamline analyses.

Applications of RNA-Seq

RNA-Seq is used in diverse research areas, including:

Cancer Transcriptomics: Identifying tumor-specific expression profiles.
Developmental Biology: Studying dynamic transcriptome changes.
Drug Discovery: Screening genes modulated by therapeutic compounds.

Conclusion

RNA-Seq analysis is a cornerstone of modern transcriptomics, offering bioinformaticians a versatile toolkit for unraveling gene expression and regulation. Mastering RNA-Seq workflows and tools empowers researchers to transform raw sequencing data into biological discoveries.

Whether you’re investigating disease mechanisms, exploring cellular pathways, or developing new therapeutics, RNA-Seq is a powerful ally in your bioinformatics arsenal.