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from GenomeWeb News

NEW YORK (GenomeWeb News) – Worker wasps have a more active transcriptome than queen wasps do, Seirian Sumner, a senior lecturer at the University of Bristol, and her colleagues reported in Genome Biology yesterday.

Workers, queens, and other social wasp castes share the same genome, but how that genome leads to alternative phenotypes like workers and queens is not fully known, especially as social wasps, and other social insects, are derived from non-social, solitary ancestors. In social wasps, queen wasps are responsibly mainly for reproduction while workers care for the offspring. Read more

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from Bio-IT World

Sales of the Ion Torrent desktop sequencers exceed those of its rival MiSeq machine from Illumina, according to a new financial report, which highlights encouraging trends for both Illumina and Ion Torrent parent company, Life Technologies.

In the DNA Sequencing report issued late last week from Macquarie Equities Research, analysts Jon Groberg and Travis Steed write that Illumina has made “great strides” with the MiSeq instrument, but “Life’s position in the desktop category remains underappreciated.”

According to Macquarie, Ion Torrent has about 62% of the emerging desktop sequencing market, with 54% the PGM, 8% the Ion Proton, which launched last year. The MiSeq currently has about 38% market share.

The report says Illumina has placed about 1,100 MiSeqs so far – about 200-250 per quarter throughout 2012 — compared to around 1,600 Ion Torrent PGMs. (The surplus is attributed to the earlier launch of the PGM.) Sales of the MiSeq equaled or exceeded PGM placements each quarter in 2012, says the report. Life Technologies also sold about 230 Ion Proton instruments last year.

Macquarie estimates Ion captured about 53% of desktop revenues (PGM 36%) last year, the MiSeq 47%. Consumable revenues are fairly even on the PGM and MiSeq, with users typically spending about $45-50,000 per instrument.

Anecdotal feedback on instrument accuracy and ease of use “continues to favor MiSeq,” the report states, but Ion’s competitiveness is due to the platform’s “relatively low acquisition and per-run costs, performance trajectory and run time, ability to be ‘good enough’ for key applications, and Life’s commercial reach.” Read more

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The lack of a miRNA set and genome sequence of wild rice (Oryza rufipogon) has prevented us from determining the role of miRNA genes in rice domestication. In this study, a team led by researchers at the Zhejiang University, China sequenced three small RNA populations and a degradome of O. rufipogon by the Illumina sequencing platform and investigated the expression levels of microRNAs (miRNAs) by miRNA chips. A de novo O. rufipogon genome was assembled using c. 55× coverage of raw sequencing data and a total of 387 miRNAs were identified in the O. rufipogon genome based on c. 5.2 million unique small RNA reads from three different tissues of O. rufipogon. Of these, O. rufipogon miRNAs, 259 were not found in the cultivated rice, suggesting a loss of these MIRNAs in the cultivated rice. They also found that 48 miRNAs were novel in the cultivated rice, suggesting that they were potential targets of domestication selection. Some miRNAs showed significant expression differences between wild and cultivated rice, suggesting that expression of miRNA could also be a target of domestication, as demonstrated for the miR164 family. Their results illustrated that miRNA genes, like protein-coding genes, might have been significantly shaped during rice domestication and could be one of the driving forces that contributed to rice domestication.

• Wang Y, Bai X, Yan C, Gui Y, Wei X, Zhu QH, Guo L, Fan L. (2012) Genomic dissection of small RNAs in wild rice (Oryza rufipogon): lessons for rice domestication. New Phytol 196(3), 914-25. [abstract]

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SAN DIEGO–(BUSINESS WIRE)–Jan. 8, 2013– Illumina (NASDAQ: ILMN) today announced a series of product and technology innovations for its powerful sequencing ecosystem – from sample preparation to system enhancements to data analysis –– that will enable the next breakthroughs in understanding the genome.

“Illumina has consistently led the market in conceptualizing, developing, and executing on industry-changing sequencing technology, and we continue to provide scientific advances that facilitate fully integrated and highly economical sequencing with very rapid turnaround,” said Jay Flatley, President and Chief Executive Officer of Illumina. “These capabilities allow us to continue to meet the evolving needs of our customers, as they develop an ever-increasing range of applications, in new and emerging markets from agrigenomics to molecular diagnostics.” Read more

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from GenomeWeb

NEW YORK (GenomeWeb News) – Two investment banks today issued bullish opinions on Illumina, noting the continued uptake of the company’s next-generation sequencers as well as the benefits of persistent rumors about a possible purchase of Illumina by Roche, and the penetration of the technology into the diagnostics arena.

Piper Jaffray upgraded the San Diego firm to an Overweight rating from Neutral and raised the price target on Illumina’s stock to $61 from $47. Separately, Leerink Swann increased its valuation to a range of $62 to $63 from an earlier range of $57 to $59 while it maintained an Outperform rating.

In a research note Piper Jaffray’s William Quirk said that its internal data indicates Illumina continues to gain market share in the next-generation sequencing market with the HiSeq installed based increasing to 730 systems to date from 645 at the end of June, a 13 percent improvement. At the same time, the installed base for MiSeq’s jumped 101 percent to 141 instruments from 70 during the same period. Read more

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Centro di Biotecnologie Molecolari, Via Nizza 52, Torino, Italy, 27-28 March 2013

Next Generation Sequencing platforms changes the conventional view of transcript analysis. Massive Parallel Sequencing methods (MPS) can extend and improve the knowledge obtained by conventional microarray technology, both for mRNAs and short non-coding RNAs, e.g. miRNAs.

This hands-on course is organized by B&Gu with guest speakers from Illumina , Singapore Immunology Network and Polytechnic of Torino. The course is suitable for biologists who are new to RNA-seq technology. Additionally, if you have analyzed gene expression data from microarrays and would now like to do projects using RNA-seq technology, then this workshop will enable you to gather information on the critical issue on RNA-seq from sample prep to data analysis.

The course is based on the use of Bioconductor open-source software. However, R coding skill is not required since all the analyses are done using oneChannelGUI, a graphical interface to Bioconductor tools, designed for life scientists who are not familiar with R language.

Knowledge of statistics is not necessary prior to attending the course.

Further course information, agenda and logistics can be found in the course booklet.

(more info…)

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from GenomeWeb

Illumina Battles BGI Over Purchase of Complete Genomics

Illumina confirmed that it made a competing unsolicited bid for Complete Genomics at about 5 percent above BGI-Shenzhen’s offer for the Mountain View, Calif.-based next-generation sequencing services firm.

In a document filed with the US Securities and Exchange Commission late Friday, Illumina confirmed it was the company identified as “Party H” by Complete Genomics that made an offer of $3.30 per share to acquire it.

Complete Genomics disclosed the bid earlier this week in an SEC document, adding that it declined the offer after determining that it would likely be turned down by regulators. In another SEC document filed on Friday, Complete Genomics further said that its board determined BGI’s offer to be pro-competitive compared to Party H’s because among other things BGI’s offer “will preserve the company’s innovative technology in the market place.”

(read more…)

Helicos BioSciences Files for Chapter 11 Bankruptcy Protection

Helicos BioSciences filed for Chapter 11 bankruptcy protection on Thursday.

The Cambridge, Mass.-based single-molecule sequencing firm filed its petition in US Bankruptcy Court for the District of Massachusetts after years of struggling financially and trying to keep pace in the fast-moving next-generation sequencing space.

Under Chapter 11 bankruptcy, Helicos will be afforded the opportunity to reorganize its operations, and the company said that it continues to operate as a debtor in possession.

In a document filed with the US Securities and Exchange Commission today, Helicos said it filed for Chapter 11 protection after its board “determined that continued operation of the company outside of bankruptcy protection is not possible due to its lack of cash resources and no available funding operations.”

(read more…)

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Researchers at Harvard University have developed a rapid, cost-effective method (FREQ-Seq) that leverages Illumina next-generation sequencing for localized, quantitative allele frequency detection. Analogous to RNA-Seq, FREQ-Seq relies upon counts from the >10⁵ reads generated per locus per time-point to determine allele frequencies. Loci of interest are directly amplified from a mixed population via two rounds of PCR using inexpensive, user-designed oligonucleotides and a bar-coded bridging primer system that can be regenerated in-house. The resulting bar-coded PCR products contain the adapters needed for Illumina sequencing, eliminating further library preparation. Read more

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from GenomeWeb News

NEW YORK – Illumina reported after the close of the market Tuesday that its third-quarter revenues increased 21 percent year over year, as the firm beat Wall Street estimates on the top and bottom line.

The San Diego-based genetic analysis tools firm reported total revenues of $285.9 million, up from $235.5 million for Q3 2011 and above analysts’ consensus estimate of $284.8 million. Its product revenues climbed to $262.4 million from $220.3 million, and its service and other revenue jumped to $23.5 million from $15.2 million. Read more

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Accelerating Biological Discoveries with Next-Generation SequencingJoin us for a one-day seminar to learn how Illumina next-generation sequencing (NGS) technology can help you uncover the answers to drive your research forward. A simple end-to-end sequencing workflow lets you go from sample to results—faster and easier than ever before. During this event Illumina experts will present an overview of NGS, discussing the basic concepts and key advantages over traditional technologies. You’ll also see talks from leading scientists presenting new discoveries made with NGS in cancer and microbiology research. From uncovering disease pathways to understanding complex microbial populations, the possibilities are limitless.See what NGS can do for your research.

Date:September 25, 2012Time: 9:00 AM – 3:00 PM Location: Texas Medical Center, Hornberger Conference Center 2151 W. Holcombe Blvd Houston, TX 77030

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How well does RNA-Seq data perform for quantitative whole gene expression analysis in the absence of a genome? This is one unanswered question facing the rapidly growing number of researchers studying non-model species. Researchers at the University of Helsinki, Finland have tried to answer that question by comparing the direct mapping of sequencing reads to predicted genes from the genome with mapping to de novo transcriptomes assembled from RNA-Seq data. Read more

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Genome-wide transcriptome analyses are routinely used to monitor tissue-, disease- and cell type-specific gene expression, but it has been technically challenging to generate expression profiles from single cells.

Now, a team of researchers led by scientists from the Ludwig Institute and the Karolinska Institute, Sweden, in collaboration with Illumina have developed a robust mRNA-Seq protocol (Smart-Seq) that is applicable down to single cell levels. Compared with existing methods, Smart-Seq has improved read coverage across transcripts, which enhances detailed analyses of alternative transcript isoforms and identification of single-nucleotide polymorphisms.

The scientists determined the sensitivity and quantitative accuracy of Smart-Seq for single-cell transcriptomics by evaluating it on total RNA dilution series.  They found that although gene expression estimates from single cells have increased noise, hundreds of differentially expressed genes could be identified using few cells per cell type. They next applied Smart-Seq to circulating tumor cells from melanomas and identified distinct gene expression patterns, including candidate biomarkers for melanoma circulating tumor cells. This protocol will be useful for addressing fundamental biological problems requiring genome-wide transcriptome profiling in rare cells.

• Ramsköld D, Luo S, Wang YC, Li R, Deng Q, Faridani OR, Daniels GA, Khrebtukova I, Loring JF, Laurent LC, Schroth GP, Sandberg R. (2012) Full-length mRNA-Seq from single-cell levels of RNA and individual circulating tumor cells. Nat Biotechnol [Epub ahead of print]. [abstract]

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• SEQanswers – RNA Sequencing

  • RNAseq (SOLiD) from 18 - 200 nt June 18, 2013
    We are interested in small non-coding RNAs. Whomever you ask about the size range of small RNAs, you get a different answer. ;) Lets assume, small... […]
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  • Unmapped ratio very high on mouse genome June 17, 2013
    Hi, My problem regards RNA-Seq data. I've downloaded public data (SAGE libs w/ 6 different samples from mouse liver ) to analyse using ArrayStudio.... […]
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  • RNASeq: Read length different from expected June 17, 2013
    Hello all, I have received paired-end reads for 40 samples. The reads are supposed to be 100bp per end. Instead, 20 of my samples are 101bp per... […]
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  • How to install xgawk June 16, 2013
    Hi, This is Shrujan, i have a problem while running RNA Sequencing QC. It shows an error that xgawk is not found. So please help me installing... […]
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  • RNA Sequencing QC Error while using with Sequence_QC.sh file June 15, 2013
    Hi, This is Shrujan kumar Madadha, I had an error while running QC for Drosophila Yukuba fastq RNA file using Sequence_QC.sh file of FASTX... […]
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  • Cuffmerge related query June 12, 2013
    I have a query regarding what samples should be merged using cuffmerge, when you have multiple phenotypes (each with replicates). Lets say my mouse... […]
    ParthavJailwala

• Biostar – RNA-Seq

  • edgeR: very low p-value and very high variance within the group of replicates. What's my problem??
    I'm using edgeR in order to perform differential expression analysis from RNA-seq experiment. I have 6 samples of tumor cell, same tumor and same treatment: 3 patient with good prognosis and 3 patient with bad prognosis. I want to compare the gene expression among the two groups. I ran the edgeR pakage like follow: x […]
  • Normalising tag count to RPKM
    Hi! I was wondering if their is a way to normalise the number of reads in a region and the RPKM of the nearest gene to that region, so that a correlation could be computed. Like the following data shows number of tags in first column and RPKM in second column Tags RPKM 15 0.14619 11 0 203 0.2259 129 10.701 300 7.0772 122 2.3234 346 10.666 77 3.117 201 16.749 […]
  • a simple question on RNA-Seq terminology
    This question may be very simple and basic, but I just need to confirm that I understand the differences among those terminologies in the RNA-Seq context. Suppose I have a sample called SLR, and it is sequenced on 5 lanes, so I have (among other output files) BAM files like L1_SLR, L2_SLR, L3_SLR, L5_SLR and L7_SLR.bam. Here, the letter "L" denotes […]
  • FInding regions of interest with minimum coverage
    Hi, I have a bam file of all my accepted hits (tophat output) and an gtf file with my genes of interest for which I am trying to find potential antisense transcripts. I would like to create a list - preferably one that can be visualized in a genome browser - that shows all genes that have antisense reads in the accepted hits.bam file provided that there are […]
  • How to remove the intronic reads before counting
    I got RNASeq data in several samples. I checked the FastQC, seems the read quality are good (Hiseq 2000). But the problem is many reads are mapped to intronic region, and the regions have no any reference exons there (Refseq, ensembl, gencode). We don't know what they are. We guess the problem happend in library preparation, the concentration was low. N […]
  • Which strand of the mRNA molecule does the sequencer output as a "read"?
    In Illumina Stranded RNA-Seq (using the dUTP method), do the final reads in the fastq files correspond to the initial molecule (that was transcribed), or to the reverse complement of the molecule? C […]