Hot off the presses! Aug 01 Nat Methods

The Aug 01 issue of the Nat Methods is now up on Pubget (About Nat Methods): if you're at a subscribing institution, just click the link in the latest link at the home page. (Note you'll only be able to get all the PDFs in the issue if your institution subscribes to Pubget.)

Latest Articles Include:

• Method of the Year 2009 voting begins
  - Nat Methods 6(8):547 (2009)
  We are now accepting nominations and collecting votes for the Method of the Year 2009.
• Software by any name
  - Nat Methods 6(8):547-548 (2009)
  Computational biologists are often tempted to avoid providing a named software implementation of their new algorithm, but resisting this temptation helps avoid difficulties later on and benefits the wider community of biologists.
• MAQGene: software to facilitate C. elegans mutant genome sequence analysis
  - Nat Methods 6(8):549 (2009)
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• SHOREmap: simultaneous mapping and mutation identification by deep sequencing
  - Nat Methods 6(8):550-551 (2009)
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• Sub-angstrom accuracy in protein loop reconstruction by robotics-inspired conformational sampling
  - Nat Methods 6(8):551-552 (2009)
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• Mass spectrometers on a chip
  - Nat Methods 6(8):555 (2009)
  A prototype for a mass spectrometer with single-molecule sensitivity has prospects for single-cell proteomics.
• Actions speak louder
  - Nat Methods 6(8):556-557 (2009)
  A fluorescence resonance energy transfer (FRET)-based biosensor helps scientists monitor the activation of an essential signaling protein over the course of embryogenesis in Drosophila melanogaster.
• New shapes of prions
  - Nat Methods 6(8):556-557 (2009)
  Modification of a system for rapid amplification of misfolded prion proteins allows de novo generation of these infectious molecules and provides a glimpse of the diverse range of possible misfolded prion strains.
• News in brief
  - Nat Methods 6(8):557 (2009)
  
• Network countdown
  - Nat Methods 6(8):558 (2009)
  Researchers lay the foundation for networks that will self-destruct after being exposed to a predetermined number of stimuli.
• Clinical proteomics on target
  - Nat Methods 6(8):560 (2009)
  A multilaboratory study designed to assess the reproducibility of multiple reaction monitoring (MRM) mass spectrometry–based proteomics demonstrates the promise of this technology for disease biomarker verification.
• A good nose for stem cells
  - Nat Methods 6(8):562 (2009)
  Cells can be delivered to the rodent brain noninvasively, via the nasal cavity.
• Probing the basis for genotype-phenotype relationships
  - Nat Methods 6(8):565-566 (2009)
  Transposon mutagenesis coupled with microarray analysis helps to rapidly generate information about changing genotype-phenotype relationships in laboratory-evolved bacteria.
• New genetic tools for cell lineage analysis in Drosophila
  - Nat Methods 6(8):566-568 (2009)
  Real-time lineage tracing in flies gets a boost with three techniques to specifically label a progenitor's daughter cells.
• Statistical methods for analysis of high-throughput RNA interference screens
  - Nat Methods 6(8):569-575 (2009)
  RNA interference (RNAi) has become a powerful technique for reverse genetics and drug discovery, and in both of these areas large-scale high-throughput RNAi screens are commonly performed. The statistical techniques used to analyze these screens are frequently borrowed directly from small-molecule screening; however, small-molecule and RNAi data characteristics differ in meaningful ways. We examine the similarities and differences between RNAi and small-molecule screens, highlighting particular characteristics of RNAi screen data that must be addressed during analysis. Additionally, we provide guidance on selection of analysis techniques in the context of a sample workflow.
• Conditional and reversible disruption of essential herpesvirus proteins
  - Nat Methods 6(8):577-579 (2009)
  Elucidating the function of essential proteins of complex pathogenic viruses is impeded by a paucity of complementing systems. By fusing a destabilizing domain of the FK506-binding protein to essential cytomegalovirus proteins, we generated virus mutants in which amounts of fusion proteins and viral growth can be regulated by the synthetic ligand shield-1. This conditional approach will greatly facilitate the analysis of gene functions of herpesviruses and viruses of other families.
• Global discovery of adaptive mutations
  - Nat Methods 6(8):581-583 (2009)
  Although modern DNA sequencing enables rapid identification of genetic variation, characterizing the phenotypic consequences of individual mutations remains a labor-intensive task. Here we describe array-based discovery of adaptive mutations (ADAM), a technology that searches an entire bacterial genome for mutations that contribute to selectable phenotypic variation between an evolved strain and its parent. We found that ADAM identified adaptive mutations in laboratory-evolved Escherichia coli strains with high sensitivity and specificity.
• Mass spectrometry of membrane transporters reveals subunit stoichiometry and interactions
  - Nat Methods 6(8):585-587 (2009)
  We describe a general mass spectrometry approach to determine subunit stoichiometry and lipid binding in intact membrane protein complexes. By exploring conditions for preserving interactions during transmission into the gas phase and for optimally stripping away detergent, by subjecting the complex to multiple collisions, we released the intact complex largely devoid of detergent. This enabled us to characterize both subunit stoichiometry and lipid binding in 4 membrane protein complexes.
• Metabolic network analysis integrated with transcript verification for sequenced genomes
  - Nat Methods 6(8):589-592 (2009)
  With sequencing of thousands of organisms completed or in progress, there is a growing need to integrate gene prediction with metabolic network analysis. Using Chlamydomonas reinhardtii as a model, we describe a systems-level methodology bridging metabolic network reconstruction with experimental verification of enzyme encoding open reading frames. Our quantitative and predictive metabolic model and its associated cloned open reading frames provide useful resources for metabolic engineering.
• Virtual terminator nucleotides for next-generation DNA sequencing
  - Nat Methods 6(8):593-595 (2009)
  We synthesized reversible terminators with tethered inhibitors for next-generation sequencing. These were efficiently incorporated with high fidelity while preventing incorporation of additional nucleotides, and we used them to sequence canine bacterial artificial chromosomes in a single-molecule system that provided even coverage for over 99% of the region sequenced. This single-molecule approach generated high-quality sequence data without the need for target amplification and thus avoided concomitant biases.
• Dereplication and de novo sequencing of nonribosomal peptides
  - Nat Methods 6(8):596-599 (2009)
  Nonribosomal peptides (NRPs) are of great pharmacological importance, but there is currently no technology for high-throughput NRP 'dereplication' and sequencing. We used multistage mass spectrometry followed by spectral alignment algorithms for sequencing of cyclic NRPs. We also developed an algorithm for comparative NRP dereplication that establishes similarities between newly isolated and previously identified similar but nonidentical NRPs, substantially reducing dereplication efforts.
• The twin spot generator for differential Drosophila lineage analysis
  - Nat Methods 6(8):600-602 (2009)
  In Drosophila melanogaster, widely used mitotic recombination–based strategies generate mosaic flies with positive readout for only one daughter cell after division. To differentially label both daughter cells, we developed the twin spot generator (TSG) technique, which through mitotic recombination generates green and red twin spots that are detectable after the first cell division as single cells. We propose wide applications of TSG to lineage and genetic mosaic studies.
• G-TRACE: rapid Gal4-based cell lineage analysis in Drosophila
  - Nat Methods 6(8):603-605 (2009)
  We combined Gal4-UAS and the FLP recombinase–FRT and fluorescent reporters to generate cell clones that provide spatial, temporal and genetic information about the origins of individual cells in Drosophila melanogaster. We named this combination the Gal4 technique for real-time and clonal expression (G-TRACE). The approach should allow for screening and the identification of real-time and lineage-traced expression patterns on a genomic scale.
• Robust, high-throughput solution structural analyses by small angle X-ray scattering (SAXS)
  - Nat Methods 6(8):606-612 (2009)
  We present an efficient pipeline enabling high-throughput analysis of protein structure in solution with small angle X-ray scattering (SAXS). Our SAXS pipeline combines automated sample handling of microliter volumes, temperature and anaerobic control, rapid data collection and data analysis, and couples structural analysis with automated archiving. We subjected 50 representative proteins, mostly from Pyrococcus furiosus, to this pipeline and found that 30 were multimeric structures in solution. SAXS analysis allowed us to distinguish aggregated and unfolded proteins, define global structural parameters and oligomeric states for most samples, identify shapes and similar structures for 25 unknown structures, and determine envelopes for 41 proteins. We believe that high-throughput SAXS is an enabling technology that may change the way that structural genomics research is done.
• Digital RNA allelotyping reveals tissue-specific and allele-specific gene expression in human
  - Nat Methods 6(8):613-618 (2009)
  We developed a digital RNA allelotyping method for quantitatively interrogating allele-specific gene expression. This method involves ultra-deep sequencing of padlock-captured single-nucleotide polymorphisms (SNPs) from the transcriptome. We characterized four cell lines established from two human subjects in the Personal Genome Project. Approximately 11–22% of the heterozygous mRNA-associated SNPs showed allele-specific expression in each cell line and 4.3–8.5% were tissue-specific, suggesting the presence of tissue-specific cis regulation. When we applied allelotyping to two pairs of sibling human embryonic stem cell lines, the sibling lines were more similar in allele-specific expression than were the genetically unrelated lines. We found that the variation of allelic ratios in gene expression among different cell lines was primarily explained by genetic variations, much more so than by specific tissue types or growth conditions. Comparison of expressed SNPs on ! the sense and antisense transcripts suggested that allelic ratios are primarily determined by cis-regulatory mechanisms on the sense transcripts.
• Cell culture: building a better matrix
  - Nat Methods 6(8):619-622 (2009)
  With the realization that cells interact extensively with their surrounding microenvironments during growth and development, the challenge for researchers has become designing three-dimensional culture systems that more closely mimic those relationships.