Tag Archives: Illumina

Whole genome sequencing a complete island

http://commons.wikimedia.org/wiki/File:Coat_of_arms_of_Iceland.svg

http://commons.wikimedia.org/wiki/File:Coat_of_arms_of_Iceland.svg

Two days ago a groundbreaking study was published in Nature Genetics: Whole genome sequencing of 2,636 Icelanders and Genotyping of 104,220 Icelanders.

The advantage of using a small population like the Icelanders for this kind of study is that there are fewer rare variants, but sometimes also a higher occurance of some of these variants.

For the study, geenomic DNA was isolated from white blood cells and subsequent sequencing was performed on GAIIx and HiSeq instruments. The resulting reads were aligned to the human reference genome (NCBI Build 36 (hg18).

Gudbjartsson et al. then examined the data from different angles. For example, they looked for geographical dependencies for specific variants or how the data can be used to learn more about phenotypes and their underlying genomic pattern. But they also report an example “how rare variants […] can be used to analyze clinical problems”. (Gudbjartsson et. al)

Since every human being has a unique genomic pattern I think studies like this are of high importance to learn more about disease related genotypes. This will help to gain confidence in the results that we get from molecular diagnostic assays for disease treatment now and in the future.

Read the complete publication here.

New Illumina Instruments

HiSeq_picsNew Year – New Innovations. Illumina directly starts off 2015 with a huge announcement: the launch of 4 new systems (GenomeWeb, 12th Jan).

Here a short overview of the new systems:

  • HiSeq X Five – scaled down version of the X Ten; costs: $6 million
  • HiSeq 3000  – uses a single flow cell and offers a lower price per data point than the HiSeq 2500; half the throughput (750G) as the HiSeq 4000; costs: $740,000
  • HiSeq 4000 – uses a dual flow cell and can sequence up to 12 genomes or 180 exomes in 3,5 days or less; costs: $900,000
  • NextSeq 550 – combines microarray scanning with NGS; applications: cytogenetics & prenatal genetic diagnostsis; costs: $275,000

Now I am curious to see if also other providers will have such surprising news as Illumina. We will keep you posted…

Whose genome has been sequenced? Brassica napus

de-novo-sequencingBrassicas napus, also known as oilseed rape, was formed more than 7000 years ago by allopolyploidy (chromosome doubling from to Brassicas species). Of course the genome mutated further and so it is known today that during this evolution some genes were preserved and further “improved” (e.g. oil biosynthesis genes), whereas others were lost over the course of time (e.g. glusoinolate genes).

Chalhoub et. al now sequenced the genome, because it can help to “provide insights into allopolyploid evolution and its relationship with crop domestication and improvement” (Chalhoub et. al).

What was sequenced?

Young fresh leaves from the Brassica napus French homzygous winter line “Darmor-bzh“.

Sequencing strategy: Whole genome sequencing

  1. Libraries & Sequencing:
    Roche GS FLX: ~ 70 Million reads, Average Read length: ~ 368 bp, Genome coverage: 21.2 %
    Sanger BAC Seq: 141k reads, Read length: 650 bp; Genome coverage: 0.1%
    Illumina HiSeq:  ~375 Million reads, Read length: 36, 76, 108 and 150 bp, Genome coverage: 53.9%
  2. Data output: 44.146 contigs and 20.702 scaffolds
  3. Results: A final assembly of 849.7 Mb (using SOAP and Newbler) with 89% nongapped sequences.

After genome assembly the genome was mapped to other species (e.g. B. rapa and B. oleracea) and this helped to find several interesting genes and gene variation that help to understand the complete evolution better.

Read the complete publication here.

Whose Genome Has Been Sequenced? – Recent posts:

Think Big: The UK 100,000 Genome Project

In late 2012 the 100,000 genome project was launched. UK Prime Minister David Cameron announced a new initiative led by the National Health Service to sequence the genomes of up to 100,000 people and to use their genomic information in treatment and studies of cancer and other diseases. The government set aside 100 million GBP for this project.

hiseq-x-tenGenomics England which is heading the project now named 10 firms that have been selected to for the assessment of the next phase of the project. The companies are Congenica; Diploid; NantOmics; Genomics Ltd.; Illumina; Qiagen; Lockheed Martin; NextCode Health; Omicia; and Personalis.

As part of the recently completed stage, Genomics England in February sent out a questionnaire to 28 participants in relation to 10 cancer/normal samples and 15 rare disease trio samples.

Illumina is partnering as well and will contribute with the ultra-high throughput sequencing platform HiSeq XTM Ten.

What will be the next step? Sequencing everyone?

More Updates: Illumina & IonTorrent

Quarter 4 of 2014 seems to be another exciting one for Next Generation Sequencing. Beside the chemistry update for PacBio RSII also Illumina and IonTorrent / ThermoFisher announced two major improvements / achievements:

  • Chemistry update for the Illumina HiSeq X Ten and the HiSeq 2500 Rapid Run
    The new v2 reagent kit for the HiSeq X Ten supports a PCR-free sample preparation kit, which eliminates amplification during the library preparation. So far only sample preparation kits with PCR were possible, which sometimes results in a lower quality of challeging genomic regions.
    The new v2 reagent kit for the HiSeq 2500 enables users to sequence 2x 250 bp and the new chemistry therefore delivers up to 300 Gbp of data in only 60 hours. (Press Release)
    To my opinion Illumina proves once more that NGS is highly dynamic and that their continous update for existing systems is the key for their success (the latest financial report confirms that Q3 of 2014 with a growth of 10% is the strongest since 2011 for Illumina (Fierce Medical Devices)).
  • IonTorrent goes diagnostic
    The Ion PGM Dx System is now also CE-Marked for in vitro diagnostic (IVD) use in Europe. Thermo Fisher Scientific believes that the CE-mark “will enable European clinical laboratories to more easily […] implement new […] diagnostic assays” (Press Release).
    In September they announced already that the PGM is now listed with the U.S. FDA as a Class II Medical Device.
    To my opinion the clearance for diagnostic use in Europe as well as in the U.S. will further strengthen the position of the Ion PGM in clinical laboratories.

Whose Genome Has Been Sequenced? Belgica antarctica

de-novo-sequencingExtreme conditions require extreme actions. And this is what the midge Belgica antarctica has done. The midge lives exclusively in the Antarctic and in order to survive shrinked its genome to the smallest possible size. As of today, this is the smallest insect genome that has been sequenced.

Kelley et. al. now sequenced the genome of Belgica antarctica with the aim to learn more about how insects in general can adapt to the most extreme conditions.

What was sequenced?

Two fourth instar larva (Belgica antarctica) collected near Palmer Station, Antarctica.

Sequencing strategy: Whole genome sequencing & RNA-sequencing

  1. Libraries & Sequencing: 1 channel 2x 100 bp Illumina HiSeq 2000 (SG library (400 bp insert)) and one SMRT-cell of a 10 kb fragment library on PacBio RSII (P4 DNA Polymerase)
  2. Data output: 92 M paired-end reads from the shotgun sequencing with Illumina. These resulted in 5,422 contigs. Using the paired-end RNA-Seq data the number of contigs has been reduced to 5,064. Genome coverage with Illumina sequencing ~ 100x.
  3. Results: The total genome is ~ 99 Mbp.

For the PacBio sequencing a second larvae was used. But due to the low input of genomic DNA the PacBio data yielded only in a modest improvement in assembly. This underlines the need of a long-read sequencing technology with low input DNA material.

The de novo sequencing of the midge Belgica antarctica revealed that the smalll genome size is achieved by a reduction in repeats, TEs and intron size.

Read the complete publication here.

Whose Genome Has Been Sequenced? – Recent posts:

Why is Illumina so successful? Watch an interview with Illuminas CEO

In the 2nd quarter of 2014 Illumina reported adjusted earnings of 57% per share – most probably the biggest increase in the companies history. Watch this interview with the CEO of Illumina, Jay Flateley, to learn more about the reasons of Illumina’s success.

 

The Common Marmoset as a Model Organism for the Study of Drug Metabolism

marmosetSeveral non-human primates including Macaca mulatta and Macaca fascicularis are well known as experimental animals in the field of neuroscience, stem cell research, drug toxicology, and other applications. The common marmoset (Callithrix jacchus) is also a non-human primate and is suitable as experimental animal because of the small size and highfecundity.

For developing a drug metabolism model, our collaborators and Eurofins Genomics (2014) performed transcriptome analysis of the common marmoset using in parallel long-read technology (Roche GS FLX+) and short-read sequencing (Illumina HiSeq 2000). This parallel NGS approach resulted in both, the identification and the quantitative analysis of transcripts and thus giving insight into gene expression during drug metabolism. Finally we obtained rich information about genes involved in drug-metabolism including 18 cytochrome P450- and 4 flavin-containing monooxygenase -like (FMO) genes, and their tissue-specific expression patterns.

The results of this study are the foundation for future studies not limited to drug metabolism & pharmacokinetics.

Is China breaking the dominance of Illumina?

BIGIS-4 is the name of an independently developed next generation sequencer made in China. The sequencer shall challenge the dominance of Illumina. On 18 April, scientists from the Beijing Institute of Genomics (BIG) of the Chinese Academy of Sciences and partner company Zixin Pharmaceutical Industrial Co Ltd demonstrated their BIGIS-4 sequencing machine in Changchun, Jilin province.

The Chinese machine has a longer read length than dominant sequencers like those made by Illumina in the US. Its manufacturing cost will be one third cheaper than imported machines, and operation costs about one fifth lower, according to Yu Jun of BIG, chief scientist of the project. Yu was also a co-founder of Shenzhen-based BGI, a spin-off of BIG and now the world’s largest sequencing service provider.

Yu’s sequencer differs from Illumina’s in that the fluorescent tag is cleaved from the newly synthesised DNA as it is incorporated, so that the reading speed is much quicker. This is similar to the pyrosequencing technology employed by Roche Diagnostics’ subsidiary 454 Life Sciences.

A publication about the complete genome sequencing and assembly of a Glaciecola mesophila spec. with BIGIS-4 is published here.

New kids on the show – who will be the winner?

Next Generation Sequencing is still a quite young market. Therefore we face the same situation every year: there is a lot of innovation going on in regard to new technologies, new instruments and other inventions. Amongst all these innovations GenomeWeb picked out the new platforms and asked in a survey about the expectancies in the market.

Instruments that were part of this survey are (at least in one of several questions):

  • Oxford Nanopore’s MinIon
  • Illumina’s X-Ten
  • Illuminas NextSeq 500
  • QIAGEN’s GeneReader
  • Life Technologies’ Ion Torrent PGM
  • Illumina HiSeq

Here are some of the results:

  • 35% of the participants say that the MinIon has the greatest impact on the sequencing community
  • 30% of the participtants will purchase most likely the NextSeq 500
  • Illumina HiSeq / MiSeq outperform the Ion Torrent Proton / PGM in data accuracy and throughput, the Ion Torrent instruments are better in respect of run time and instrument price

Read the complete survey here.