demo bam file Ion Torrent 314 chip of E. coli 400 bp run for download

BAM file of B22-730 (314v2 E. coli 400 bp run)
Ion Torrent PGM 314v2 run with a mode read length of 400bp and per-base raw read accuracy >99%.

https://s3.amazonaws.com/ion-torrent/pgm/B22-730/B22-730.bam

Source: https://apps.thermofisher.com/apps/publiclib/#/datasets

Freelancing in Bioinformatics? It's happening here...uBiome FASTQ

http://www.guru.com/jobs/ubiome-raw-data-fastq-files-analysis/1210516#proposalModal

Any takers to help walk this guy through analysing ubiome raw fastq files?

p.s. I wasn't aware that ubiome gives out fastq files?

Illumina produces 3k of 8500 bp reads on HiSeq using Moleculo Technology

Keith blogged about how super long read sequencing methods would be a threat to Illumina in Jan 2013. Today, Illumina can now openly acknowledge the shortcomings of their short reads for various applications like

• assembly of complex genomes (polyploid, containing excessive long repeat regions, etc.), 
• accurate transcript assembly, 
• metagenomics of complex communities, 
• and phasing of long haplotype blocks.

the reason?
This latest set of data released on BaseSpace

Read length distribution of synthetic long reads for a D. melanogaster library

The data set, available as a single project in BaseSpace, can be accessed here.

image source: http://blog.basespace.illumina.com/2013/07/22/first-data-set-from-fasttrack-long-reads-early-access-service/

with the integration of Moleculo they have managed to generate ~30 gb of raw sequence data. They have refrained from talking about 'key analysis metrics' that's available in the pdf report. Perhaps it's much easier to let the blogosphere and data scientists dissect the new data themselves.

Am wondering when the 454 versus Illumina Long Reads side-by-side comparison will pop up

UPDATE:

Can't find the 'key analysis metrics' in the pdf report files. Perhaps it's still being uploaded? *shrugs*
so please update me if you see it  otherwise I just have to run something on it

These are the files that I have now

total 512M
 259M Jul 18 01:01 mol-32-2832.fastq.gz
  44K Jul 24  2013 FastTrackLongReads_dmelanogaster_281c.pdf
 149K Jul 24  2013 mol-32-281c-scaffolds.txt
  44K Jul 24  2013 FastTrackLongReads_dmelanogaster_2832.pdf
 151K Jul 24  2013 mol-32-2832-scaffolds.txt
 253M Jul 24  2013 mol-32-281c.fastq.gz

md5sums
6845fc3a4da9f93efc3a52f288e2d7a0  FastTrackLongReads_dmelanogaster_281c.pdf
02f5de4f7e15bbcd96ada6e78f659fdb  FastTrackLongReads_dmelanogaster_2832.pdf
586599bb7fca3c20ba82a82921e8ba3f  mol-32-281c-scaffolds.txt
b25010e9e5e13dc7befc43b5dff8c3d6  mol-32-281c.fastq.gz
6822cfbd3eb2a535a38a5022c1d3c336  mol-32-2832-scaffolds.txt
873f09080cdf59ed37b3676cddcbe26f  mol-32-2832.fastq.gz

I have ran FastQC (FastQC v0.10.1) on both samples the images below are from 281c.
you can download the full HTML report here
https://www.dropbox.com/sh/5unu3zba9u21ywj/JT4HdkzfOP/mol-32-281c_fastqc.zip
https://www.dropbox.com/s/mpxa5wx51iqmiz3/mol-32-2832_fastqc.zip

Reading about the Moleculo sample prep method, it seems like it's just a rather ingenious way to stitch short reads which are barcoded to form a single long contig. if that is the case, then I am not sure if the base quality scores here are meaningful anymore since it's a mini-assembly. Also this takes out any quantitative value of the number of reads I presume. So accurate quantification of long RNA molecules or splice variants isn't possible. Nevertheless it's an interesting development on the Illumina platform. Looking forward to seeing more news about it.

Other links

Illumina Long-Read Sequencing Service
Moleculo technology: synthetic long reads for genome phasing, de novo sequencing
CoreGenomics: Genome partitioning: my moleculo-esque idea
Moleculo and Haplotype Phasing - The Next Generation TechnologistNext Generation Technologist
Abstract: Production Of Long (1.5kb – 15.0kb), Accurate, DNA Sequencing Reads Using An Illumina HiSeq2000 To Support De Novo Assembly Of The Blue Catfish Genome (Plant and Animal Genome XXI Conference)
http://www.moleculo.com/ (no info on this page though)
Illumina Announces Phasing Analysis Service for Human Whole-Genome Sequencing - MarketWatch

Illumina Announces Moleculo Long Read Technology and Phasing As Service
First publication using the Long Read Seq (LRseq) The genome sequence of the colonial chordate, Botryllus schlosseri | eLife Contains a diagram explaining the LRSeq protocol. This experiment yielded ~1000 6.3kb fragments

Patent information on the Long Read technology
https://docs.google.com/viewer?url=patentimages.storage.googleapis.com/pdfs/US20130079231.pdf

BMC Genomics. 2011 Sep 7;12(1):444. [Epub ahead of print]

WebMGA: a Customizable Web Server for Fast Metagenomic Sequence Analysis.

Wu S, Zhu Z, Fu L, Niu B, Li W.

Abstract

ABSTRACT:

BACKGROUND:

The new field of metagenomics studies microorganism communities by culture-independent sequencing. With the advances in next-generation sequencing techniques, researchers are facing tremendous challenges in metagenomic data analysis due to huge quantity and high complexity of sequence data. Analyzing large datasets is extremely time-consuming; also metagenomic annotation involves a wide range of computational tools, which are difficult to be installed and maintained by common users. The tools provided by the few available web servers are also limited and have various constraints such as login requirement, long waiting time, inability to configure pipelines etc.

RESULTS:

We developed WebMGA, a customizable web server for fast metagenomic analysis. WebMGA includes over 20 commonly used tools such as ORF calling, sequence clustering, quality control of raw reads, removal of sequencing artifacts and contaminations, taxonomic analysis, functional annotation etc. WebMGA provides users with rapid metagenomic data analysis using fast and effective tools, which have been implemented to run in parallel on our local computer cluster. Users can access WebMGA through web browsers or programming scripts to perform individual analysis or to configure and run customized pipelines. WebMGA is freely available at http://weizhongli-lab.org/metagenomic-analysis.

CONCLUSIONS:

WebMGA offers to researchers many fast and unique tools and great flexibility for complex metagenomic data analysis.

PMID:

21899761

[PubMed - as supplied by publisher]

Chimeric 16S rRNA sequence formation and detection in Sanger and 454-pyrosequenced PCR amplicons [RESOURCES]

Chimeric 16S rRNA sequence formation and detection in Sanger and 454-pyrosequenced PCR amplicons [RESOURCES]

from Genome Research current issue by Haas, B. J., Gevers, D., Earl, A. M., Feldgarden, M., Ward, D. V., Giannoukos, G., Ciulla, D., Tabbaa, D., Highlander, S. K., Sodergren, E., Methe, B., DeSantis, T. Z., The Human Microbiome Consortium, Petrosino, J. F., Knight, R., Birren, B. W.

Bacterial diversity among environmental samples is commonly assessed with PCR-amplified 16S rRNA gene (16S) sequences. Perceived diversity, however, can be influenced by sample preparation, primer selection, and formation of chimeric 16S amplification products. Chimeras are hybrid products between multiple parent sequences that can be falsely interpreted as novel organisms, thus inflating apparent diversity. We developed a new chimera detection tool called Chimera Slayer (CS). CS detects chimeras with greater sensitivity than previous methods, performs well on short sequences such as those produced by the 454 Life Sciences (Roche) Genome Sequencer, and can scale to large data sets. By benchmarking CS performance against sequences derived from a controlled DNA mixture of known organisms and a simulated chimera set, we provide insights into the factors that affect chimera formation such as sequence abundance, the extent of similarity between 16S genes, and PCR conditions. Chimeras were found to reproducibly form among independent amplifications and contributed to false perceptions of sample diversity and the false identification of novel taxa, with less-abundant species exhibiting chimera rates exceeding 70%. Shotgun metagenomic sequences of our mock community appear to be devoid of 16S chimeras, supporting a role for shotgun metagenomics in validating novel organisms discovered in targeted sequence surveys.

JCVI Supports Human Mircrobiome Body Site Experts with Shotgun Data Analysis from JCVI Blog by Johannes Goll

The current survey comprises more than 700 samples from hundreds of individuals taken from up to 16 distinct body sites. Illumina sequencing has yielded more than 20 billion Illumina reads and annotation data produced from the sequences exceeds 10 terabytes. In anticipation of such data volumes, we have developed JCVI Metagenomics Reports (METAREP), an open source tool for high-performance comparative analysis, in 2010. The tool enables users to slice and dice data using a combination of taxonomic and functional/pathway signatures. To demonstrate how the tool can be used by body site experts, we picked and loaded sample data from 17 oral samples and presented a quick tutorial on how users can view, search, browse individual samples and compare multiple samples (see video). The functionality was very well received and body site experts asked JCVI to make all the 700+ samples available. As a result of the Jamboree, JCVI in agreement/collaboration with the HMP Data Analysis and Coordination Center and the rest of the HMP consortium, will soon set-up a dedicated HMP METAREP instance that will allow body-site experts and eventually other users to analyze the DAWG data in a user-friendly way via the web.

Microbial community profiling workshop in UK

Mike Head left a comment about a workshop which looks interesting. Would loved to be there.

http://idrn.org/events/upcoming/microbial.php

AIM
The aim of such an event would be to bring together researchers interested in profiling whole bacterial communities using techniques such as metagenomics, (16s rDNA sequencing and whole bacterial DNA sequencing), T-RFLP, and other commercial applications.

OBJECTIVES
To allow interested researchers and clinicians to better understand the potential of such techniques and to identify the best method of profiling their bacterial community of choice. To provide researchers who wish to use such methods with an overview of the theory of these methods such that experimental design is possible. Use of methods will be illustrated with examples of their use in infection research

IMPORTANCE Such techniques are used in environmental microbiology and are now being used in projects such as the human microbiome projects to investigate the diversity human pathogens and commensals and to characterise the microbial profile of various niches and relate this to conditions such as GI and respiratory disease.

CONFIRMED SPEAKERS
Alan Walker (Sanger, UK)
Geraint Rogers (Kings, UK)
Mike Cox (Imperial, UK)
Bill Cookson (Imperial, UK)
Nick Loman / Mark Pallen (Bham, UK)
Chris Quince (Glasgow, UK)
Luanne Hall-Stoodley (Southampton, UK)
Bill Keevil (Southampton, UK)
Nick Jakubovics (Newcastle, UK)

How much data for Metagenomics?

Interesting read for bioinformaticians doing metagenomics (the title says it all).
Within it, it asks the very pertinent question of How much data? However, the suggested data size of 100 Mbp is for Sanger Sequencing. Any readers with comments on the suggested data size for NGS?


How Much Sequence Data?
A common question asked by researchers embarking on their first metagenomic analysis is how much sequence data they should request or allocate for their project. Unlike genome projects, metagenomes have no fixed end point, i.e., a completed genome.......For example, if a dominant population represents 10% of the total community and 100 Mbp is obtained, then this population is expected to be represented by 10 Mbp, assuming completely random sampling of the community. If the average genome size of individuals in this population is 2 Mbp, then an average of 5x coverage of the composite population genome will be expected. To place this in perspective, 6x to 8x coverage of microbial isolates is a common target to obtain a draft genome suitable for finishing.

METAREP is a new open source tool developed for high-performance comparative metagenomics

Found this blog post at JCVI
Are your carrying out large scale metagenomics analyses to identify differences among multiple sample sites? Are you looking for suitable analysis  tools?
If you have not yet found the right analysis tool, you may be interested in  the latest beta version of JCVI Metagenomics Reports (METAREP)  [Test It].
METAREP is a new open source tool developed for high-performance comparative metagenomics .
It provides a suite of web based tools to help scientists view, query, browse, and compare metagenomics annotation data derived from ORFs called on metagenomics reads or assemblies.

PyroNoise:Accurate determination of microbial diversity from 454 pyrosequencing data

Using 454 to do microbial ecology / metagenomics of environmental / soil samples?
Then I think you should take a look at this paper.

Quince, C., Lanzén, A., Curtis, T., Davenport, R., Hall, N., Head, I., Read, L., & Sloan, W. (2009). Accurate determination of microbial diversity from 454 pyrosequencing data Nature Methods, 6 (9), 639-641 DOI: 10.1038/nmeth.1361

The Pathogens blog has a good summary post on it.

WebCARMA for metagenomic reads

Fascinating software! Will explore in time 

BMC Bioinformatics. 2009 Dec 18;10(1):430. [Epub ahead of print]

WebCARMA: a web application for the functional and taxonomic classification of unassembled metagenomic reads.

Gerlach W, Junemann S, Tille F, Goesmann A, Stoye J.

ABSTRACT: BACKGROUND: Metagenomics is a new field of research on natural microbial communities. High-throughput sequencing techniques like 454 or Solexa-Illumina promise new possibilities as they are able to produce huge amounts of data in much shorter time and with less efforts and costs than the traditional Sanger technique. But the data produced comes in even shorter reads (35-100 basepairs with Illumina, 100-500 basepairs with 454-sequencing). CARMA is a new software pipeline for the characterisation of species composition and the genetic potential of microbial samples using short, unassembled reads. RESULTS: In this paper, we introduce WebCARMA, a refined version of CARMA available as a web application for the taxonomic and functional classification of unassembled (ultra-)short reads from metagenomic communities. In addition, we have analysed the applicability of ultra-short reads in metagenomics. CONCLUSIONS: We show that unassembled reads as short as 35 bp can be used for the taxonomic classification of a metagenome. The web application is freely available at http://webcarma.cebitec.uni-bielefeld.de.

PMID: 20021646 [PubMed - as supplied by publisher]