Jive SBS Syndication Feed

Everyone Needs Next Generation Sequencing

solidcommunity@lifetech.com — Tue, 08 May 2012 16:39:27 GMT

A recent publication - Mutations in the chromatin modifier gene KANSL1 cause the 17q21.31 microdeletion syndrome, (Koolen, D et al, 2012, Nature Genetics, doi:10.1038/ng.2262), really got me excited as it emphasizes the widespread use of next generation sequencing. This paper uses a wide variety of techniques, including arrays for CNV, MLPA (multiplex ligation-dependent probe amplification) to confirm deletions, Sanger sequencing to confirm heterozygous de novo mutations and whole transcriptome sequencing using the 5500xl Genetic Analyzer to evaluate the effects on expression. This shows how next generation sequencing is now a standard tool used in an increasing number of experimental workflows and has become an absolute need for all genomics labs and increasingly all biological labs.

For those interested in the paper here are the highlights:

The 17q21.31 microdeletion syndrome, which causes facial dysmorphology and mental impairment, usually encompasses a deletion of five known genes. In this study the authors investigated four individuals who did not demonstrate the classical deletion.
A variety of tools were used to determined that two individuals had deletions of only parts of two genes, whilst two had de novo mutations in KANSL1 (nonsense mutation in one and splice variant in other); this data was enough to pinpoint KANSL1
The 5500xl Genetic Analyzer was used to carry out whole transcriptome sequencing of EBV transformed cell lines. When cells transformed with the classic microdeletion were compared to cells transformed with the normal control gene, 321 genes were identified as differentially expressed (levels of KANSL1 were reduced by half in the cell lines transformed with the 17q21.31 deletion compared to the control gene). Over 5000 differentially expressed genes were found between the cells transformed with the control vs. the individual splice variant gene. A set of 143 genes (twice expected) with enrichment for genes involved in neuronal processes were found to be shared between cells transformed with the splice variant gene and the gene with microdeletions.
Using a Drosophila model the authors were able to show that KANSL1 homolog affects regulation of complex brain function.
In conclusion the authors showed that a single gene KANSL1 with even a single de novo heterozygous mutation is enough to replicate the effects of the classical microdeletion.

AGBT 2012 posters featuring 5500 W Series Genetic Analyzers

solidcommunity@lifetech.com — Sun, 22 Apr 2012 23:42:19 GMT

If you weren't at AGBT, then you didn't get a change to see this poster.

"Accurate detection of de novo mutations in rare and common neurodevelopmental disorders" was presented by collaborators from Radboud University Nijmegen Medical Centre. The exomes were sequenced on the 5500xl W.

The poster is attached below.

LifeScope™ Genomic Analysis Software 2.5.1

solidcommunity@lifetech.com — Sun, 15 Apr 2012 20:45:53 GMT

The new version of LifeScope™ Genomic Analysis Software has arrived. Here is what’s new:

The license file, server and key has been removed to enable a simpler install process. Customers must upgrade to LifeScope 2.5.1 to remove license file requirement.
Wildfire data compatible. It takes approximately 2X the time to process 4X the data with LifeScope hardware specs.
VCF format output available for SNP analysis and indel analysis.
Several BAM Stat issues have been addressed.

Downloading software has been made simpler for customers. The latest version is available here, under DNA Sequencing, look for LifeScope.

Automation of whole transcriptome library preparation

solidcommunity@lifetech.com — Tue, 20 Dec 2011 19:56:54 GMT

Life Technologies is committed to supporting your research and continuing to simplify workflows by expanding the menu for the AB Library Builder™ System. Today we are introducing the Whole Transcriptome Core Kit for 5500 Genetic Analysis Systems (PN 4472690) that with the AB Library Builder™ System can generate 13 whole transcriptome libraries, compatible with the SOLiD® 4 System and 5500 Series Genetic Analysis Systems in a single run with reduced hands-on time.

Don’t forget you can also prep DNA fragment libraries on the AB Library Builder™ System with kits for SOLiD® 4 System and 5500 Series Genetic Analysis Systems or using the protocol on the Ion Community for Ion Personal Genome Machine™ sequencer.

Ask your sales representative for a quote for the AB Library Builder™ System at the new reduced price.

5500 Series ICS v1.2 and new sequencing kits offer > 15% improvement in throughput for long read runs

solidcommunity@lifetech.com — Thu, 15 Dec 2011 18:01:42 GMT

The 5500 Series Genetic Analyzer ICS v1.2 software is here and offers:

Greater than 15% improvement in throughput with new fwd sequencing kit on long reads (2x60 bp and 1x75 bp)
Improved reagents delivery and tracking
WFA and bead quantitation
Reports for bead assessment and barcodes
Instant abort and pre-run check list features

This software update in now available and customers can download the software by visiting www.appliedbiosystems.com/5500ics or this link.

Additional documentation is available on the 5500 Series User Hub.

Henk den Bakker from Cornell University talks to us about Salmonella enterica

solidcommunity@lifetech.com — Tue, 22 Nov 2011 18:01:16 GMT

A Cornell University food scientist has co-authored a paper titled, "Genome sequencing reveals diversification of virulence factor content and possible host adaptation in distinct subpopulations of Salmonella" which focuses on research that will help accelerate the identification of Salmonella outbreak strains. As this research is well-timed given recent stories in the news, we talked to Henk den Bakker, the publication's lead author from Martin Wiedmann's lab, to get more information about the study.

Read the full publication

Dr. Hank den Bakker:

The paper describes how you used various methods to describe differences between S. enterica species and subclades. Why is an understanding of the population structure important for human health?

In the case of S. enterica there seems to be a strong correlation between population structure, that is groups of closely related strains, and the distribution of virulence factors such as gene clusters involved in attachment to host tissues, gene clusters involved in the metabolism of host specific carbon sources and cytotoxins. Now that we know that these subgroups differ in virulence factor content we can start to look for phenotypes of these two subgroups, first with in vitro studies in tissue culture, followed by studies in animal models, and subsequently look for specific clinical presentations that are potentially associated with infections of Salmonella of these different subclades. A better understanding of the relation between Salmonella subgroups and the types of disease they cause will provide a better understanding of how these subgroups impact human health and the food industry.

Could you describe why whole genome sequencing was needed to reach your conclusions?

By choosing whole genome sequencing combined with de novo assembly of the genomes we were able to get unique information that allowed us to get the ‘whole picture’. We used the sequence information of a large number of genes of these de novo assemblies to infer the population structure and at the same compare these genomes for the presence/absence of all the genes in the complete genome, which makes it possible to see which genes are present or absent in certain subpopulations. This is in contrast to reference based assemblies, which can help to get a detailed picture of the population structure based on SNP data, but lacks the ability to discover novel genes that are not in the reference genome. With other methods (e.g., serotyping, PFGE, MLST, MLVA, microarray) we never could have obtained this amount of information.

What evidence did you see that some S. enterica serovars are adapted to human hosts?

We currently have no evidence that certain serovars (except for Typhi and Paratyphi A) are more adapted to the human host, however we now know that certain virulence factors that are found in known ‘human restricted’ serovars such as Typhi and Paratyphi A are found in a much wider variety of serovars. Future research is needed to determine if and how these virulence factors may affect the ability of these serovars to interact with human hosts.

What did you learn about virulence factors and are there any practical implications from it?

One of the things we learned about the evolution of virulence factors in this study was that certain virulence factors, that were assumed to be specific for S. Typhi, were possibly already present in the most recent common ancestor of S. enterica and were very unlikely to be acquired by recent horizontal gene transfer. We also learned that the two clades of Salmonella differ in some virulence factors, and this may have implications for clinical symptoms associated with disease caused by Salmonella enterica.

People will be want to learn about S. Typhi and its relationships to other S. enterica. Does that explain any of its history and how it originated.

One of the things we established in this study was a good list of candidates of the serovars that are most closely related to S. Typhi. Research on pathogenicity and the evolutionary origin of S. Typhi has mainly focused on comparisons of various strains representing S. Typhi and a distantly related serovar, S. Typhimurium. Comparative genomic research of S. Typhi with a wider range of Salmonella serovars, allowed us to identify its nearest relatives. Further comparative analysis of S. Typhi with its closely related serovars will give us a more precise picture of how S. Typhi evolved into the severe human pathogen it is now.

Looking for 5500 Series example data sets?

solidcommunity@lifetech.com — Wed, 16 Nov 2011 23:46:05 GMT

You can find these data sets on the SOLID software community.

5500 HuRef Mate-paired Data contains one FlowChip of sequence (60 bp x 60 bp) from a HuRef mate-paired library.
5500 Whole Transcriptome Paired-end Data contains paired-end sequencing (75bp x 35 bp) data from a MCF7 whole transcriptome library (in a single lane) and another lane of data from a pool of barcoded whole transcriptome libraries from Ambion Brain (HBR), Universal Human Reference (UHR), Hela and Ambion Placenta. These have the ERCC controls as well.

LifeScope(TM) software 2.5 is new with a Low Frequency Detection Workflow

solidcommunity@lifetech.com — Tue, 15 Nov 2011 22:30:59 GMT

LifeScope(TM) software 2.5 is now available. Here are two main features of the software update.

Low frequency variant detection workflow

The low frequency variant detection workflow uses the diBayes algorithm (with highly optimized parameters) to reveal SNPs present in a targeted resequencing experiment. The detection range can range from 1%-5% based on experimental conditions, acceptable sensitivity and specificity, and sequencing coverage. This workflow is ideal for detecting low frequency variants in cancer-related research projects.

(check out this video to hear more: http://solid.community.appliedbiosystems.com/videos/1160)

Processing time improvements

Release 2.5 has general speed improvements. In particular, the processing time required for small indel runs is improved, due to parallelization within the module. Processing time comparisons with the 2.0 release are shown in the following table:

Application type	Data description	Savings compared to LifeScope 2.0 (hours)
Whole genome mate-pair resequencing	600M reads	~5.5
Targeted resequencing paired-end	142M reads, 50x25	~0.5
Whole transcriptome paired-end	650M, indexed	~9
Small RNA	125M reads, indexed	~1

In addtion, feature requests and bug fixes have been incorporated based on the feedback from our customers.

To learn more, visit http://www.lifetechnologies.com/lifescope

Join Life Technologies & Ion Torrent at AMP

solidcommunity@lifetech.com — Fri, 11 Nov 2011 23:41:25 GMT

Join Life Technologies & Ion Torrent at the Association for Molecular Pathology (AMP) 2011 Annual Meeting

November 17–19, 2011. Grapevine, TX.

Learn more and register for workshops

From translational research to molecular testing— Life Technologies provides innovative technology and services to accomplish your genetic analysis objectives.

Attend our special corporate workshops to learn how on

Wednesday, November 16 in Grapevine C.

Next-Generation Technology for Molecular Assays

11:00 a.m.–12:00 p.m.

Presented by: Marilyn Li, MD, Baylor College of Medicine and David Keys and Jim Torrence, Life Technologies

Ion Torrent™—Translating Genomics into the Clinic

12:00–1:00 p.m.

Presented by: Chris Corless, MD, PhD, Oregon Health Sciences University

Shaping the Future of Clinical Sequencing— Advances in Exome and Multiplex Sequencing

1:00–2:00 p.m.

Presented by: Toumy Guettouche, PhD, University of Miami

script launch xsqtools

jacobnix@gmail.com — Fri, 04 Nov 2011 18:44:25 GMT

Follow this unofficial tutorial step by step to run xsq-tools on CentOS Linux 6 and OpenSuse Linux

1. If you are using CentOS Linux 6, OpenSuse Linux check which GNU LIBC version you have installed. you can use this command

[jacob@localhost xsq-tools]$ ldd – -version
ldd (GNU libc) 2.12
Copyright (C) 2010 Free Software Foundation, Inc.
This is free software; see the source for copying conditions. There is NO
warranty; not even for MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE.
Written by Roland McGrath and Ulrich Drepper.

to run xsq-tools you will need GNU libc version 2.5+ !!

2. Get Official xsq-tools software package distribution from LifeTech/ABI

3. Open your favorite Linux console command and untar/unzip xsq-tools package.

4. If you want to convert your xsq solid 5500 output files you will need to use the convertFromXSQ.sh shell script.

5. Backup the official convertFromXSQ.sh shell script.

6. Download unofficial convertFromXSQ.sh shell script and save in your official xsq-tools directory,

7. Download script from here

8. Rename script convertFromXSQ.sh1.txt to convertFromXSQ.sh

9. Set execution permission to shell script ( chmod +x convertFromXSQ.sh )

10. Change to xsq-tools directory using linux command line (cd XSQ_Tools)

11. Extras: You can run unofficial convertFromXSQ.sh shell script like this samples

$ remember, cd XSQ-Tools first

(A) $ ./convertFromXSQ.sh -c data/Frag.xsq

(B, from any location)

$ convertFromXSQ.sh -c /home/jacob/XSQ-Tools/data/Frag.xsq -o $HOME

That’s all.

Notes:

[1] If you do not set XSQ_TOOLS environment variable, the unofficial convertFromXSQ.sh set XSQ_TOOLS environment variable automatically.

[2] Follow LifeTech/ABI official readme documentation to install xsq-tools, the unofficial script updates your .bashrc ,bash_profile in user home directories but I disabled exporting PATH environment variable because I need to do some tests, if you want to run unofficial convertFromXSQ.sh from any location you will need to update and export your PATH environment variable manually.

I hope that help you , it works for me !!

You can have your exome and custom too!

solidcommunity@lifetech.com — Fri, 28 Oct 2011 18:04:43 GMT

The same high-density probe design and economical pre-capture sample multiplexing solution in our TargetSeq™ Exome Enrichment Kits are now available with custom content enabling enrichment of target regions for resequencing on the 5500 Series Genetic Analyzers, the SOLiD® System. The TargetSeq™ Custom Enrichment Kits come in four target sizes:

100kb-500kb
500kb-2Mb
2Mb-10Mb
>10Mb

Select your regions of interest by submitting a gene list or genomic coordinates, and specific probe pools for your targets will be designed using optimized algorithms.

Economical, targeted resequencing is yours with products for exome and custom content for either the 5500 Genetic Analysis or SOLiD® Systems. TargetSeq™ Custom Enrichment Kits are also available for the Ion Personal Genome Machine™ sequencer.

Ready, Set, Sequence!

Call for AGBT 2012 Abstracts

solidcommunity@lifetech.com — Wed, 26 Oct 2011 17:27:27 GMT

We want to see the SOLiD Community in Marco Island Florida for AGBT in February. The deadline for submitting abstracts is Friday Oct 28^th, so don’t delay. You can learn more at agbt.org/abstracts.html .

Submit your abstract via email to papers@agbt.org by Friday October 28, 2011.

Hope to see you in sunny Florida come February.

Reduced 5500 Series Run Times with ICS v1.1.1

solidcommunity@lifetech.com — Tue, 25 Oct 2011 00:06:26 GMT

Mission: Reduce run times

Solution: Customer installable ICS v1.1.1 for 5500 Series Genetic Analyzers

Have you heard the news? New, optimized 5500 scan times are available with the customer installable ICS version 1.1.1 for the 5500 Series Genetic Analyzers. Don’t delay! Read about how you can download ICS v1.1.1 now on the 5500 Series User Hub. Provide your instrument serial number and download the software here.

New Hybrid Assembly Stand-alone Tool Now Available

solidcommunity@lifetech.com — Sun, 23 Oct 2011 22:38:33 GMT

Have you ever wanted to perform de novo assembly using SOLiD type data? Now you can with HAPS.

HAPS (Hybrid Assembly Pipeline with SOLiD reads) is intended to assist customers to do de novo assembly using second-generation sequencing reads in a high-level manner. HAPS utilizes several modules with the functionalities of pre-processing, mapping, pairing, scaffolding, and gap-filling respectively. In other words, HAPS is a modularized scaffolder whose modules can be easily substituted so long as the Application Program Interface (API) specification is followed.

Figure 1: Diagram showing the entire hybrid assembly workflow including HAPS

A typical hybrid assembly procedure using HAPS includes several stages:

1. An assembler (e.g. Velvet or others) is used to assemble various NGS (e.g., SOLiD, Ion Torrent, 454) reads or Sanger reads into contigs in FASTA format.

2. A mapping module is used to map SOLiD mate-pair (MP) reads onto these contigs.

3.A scaffolding module is then used to build scaffolds from the contigs using the mate-pair mapping results and finally a gap-filling module is used to fill intra-scaffold gaps.

The HAPS tool is available from http://solidsoftwaretools.com/gf/project/haps/ .

NOTE: This tool is not supported by Life Technologies. Any questions, issues, problems, bugs etc. must be submitted through the solidsoftwaretools.com developers website.

SOLiD™ EZ Bead™ System Update Saves You Time

solidcommunity@lifetech.com — Wed, 19 Oct 2011 18:03:45 GMT

The SOLiD™ EZ Bead ™ System Software v1.9 update has been released. This customer installable software update includes:

Improved robustness and streamlined Enricher runs save you 30 minutes
New Amplifier run selection for templates less than 200bp
Updates for all EZ Bead™ instruments with the latest scale run selections

Initiate your software update request by sending an email to EZBeadAssist@lifetech.com