Citations

348 Citations Found

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Precision and accuracy are values we’ve held as the core of our business since starting in 1934. Delivering robotic solutions to the academic and industrial markets, our accuracy is relied upon to support the research and work that our customers do! The newest addition to our portfolio is PIXL, built to be the most accurate colony picker on the market - in the world! In defining efficiency we want to be really clear about what we mean; accuracy is picking the correct colony and it growing successfully. Some refer to this as picking efficiency, so we will too! What would this be without some data to back it up?

https://www.nature.com/articles/nature12904

Frank W. Albert, Sebastian Treusch, Arthur H. Shockley, Joshua S. Bloom & Leonid Kruglyak Nature volume 506, pages 494–497 (27 February 2014) Variation among individuals arises in part from differences in DNA sequences, but the genetic basis for variation in most traits, including common diseases, remains only partly understood. Many DNA variants influence phenotypes by altering the expression level of one or several genes. The effects of such variants can be detected as expression quantitative trait loci (eQTL)1. Traditional eQTL mapping requires large-scale genotype and gene expression data for each individual in the study sample, which limits sample sizes to hundreds of individuals in both humans and model organisms and reduces statistical power2,3,4,5,6. Consequently, many eQTL are probably missed, especially those with smaller effects7. Furthermore, most studies use messenger RNA rather than protein abundance as the measure of gene expression. Studies that have used mass-spectrometry proteomics8,9,10,11,12,13 reported unexpected differences between eQTL and protein QTL (pQTL) for the same genes9,10, but these studies have been even more limited in scope. Here we introduce a powerful method for identifying genetic loci that influence protein expression in the yeast Saccharomyces cerevisiae. We measure single-cell protein abundance through the use of green fluorescent protein tags in very large populations of genetically variable cells, and use pooled sequencing to compare allele frequencies across the genome in thousands of individuals with high versus low protein abundance. We applied this method to 160 genes and detected many more loci per gene than previous studies. We also observed closer correspondence between loci that influence protein abundance and loci that influence mRNA abundance of a given gene. Most loci that we detected were clustered in ‘hotspots’ that influence multiple proteins, and some hotspots were found to influence more than half of the proteins that we examined. The variants that underlie these hotspots have profound effects on the gene regulatory network and provide insights into genetic variation in cell physiology between yeast strains.

https://www.molbiolcell.org/doi/10.1091/mbc.e15-07-0525

Anton Khmelinskii, Matthias Meurer, Chi-Ting Ho, Birgit Besenbeck, Julia Füller, Marius K. Lemberg, Bernd Bukau, Axel Mogk, and Michael Knop Mol Biol Cell. 2016 Jan 15;27(2):360-70. doi: 10.1091/mbc.E15-07-0525. Epub 2015 Nov 25. Tandem fluorescent protein timers (tFTs) report on protein age through time-dependent change in color, which can be exploited to study protein turnover and trafficking. Each tFT, composed of two fluorescent proteins (FPs) that differ in maturation kinetics, is suited to follow protein dynamics within a specific time range determined by the maturation rates of both FPs. So far, tFTs have been constructed by combining slower-maturing red fluorescent proteins (redFPs) with the faster-maturing superfolder green fluorescent protein (sfGFP). Toward a comprehensive characterization of tFTs, we compare here tFTs composed of different faster-maturing green fluorescent proteins (greenFPs) while keeping the slower-maturing redFP constant (mCherry). Our results indicate that the greenFP maturation kinetics influences the time range of a tFT. Moreover, we observe that commonly used greenFPs can partially withstand proteasomal degradation due to the stability of the FP fold, which results in accumulation of tFT fragments in the cell. Depending on the order of FPs in the timer, incomplete proteasomal degradation either shifts the time range of the tFT toward slower time scales or precludes its use for measurements of protein turnover. We identify greenFPs that are efficiently degraded by the proteasome and provide simple guidelines for the design of new tFTs.

https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0180814

Francesco Mezzetti, Justin C. Fay, Paolo Giudici, Luciana De Vero July 6, 2017 https://doi.org/10.1371/journal.pone.0180814 Glutathione (GSH) production during wine fermentation is a desirable trait as it can limit must and wine oxidation and protect various aromatic compounds. UMCC 2581 is a Saccharomyces cerevisiae wine strain with enhanced GSH content at the end of wine fermentation. This strain was previously derived by selection for molybdate resistance following a sexual cycle of UMCC 855 using an evolution-based strategy. In this study, we examined genetic and gene expression changes associated with the derivation of UMCC 2581. For genetic analysis we sporulated the diploid UMCC 855 parental strain and found four phenotype classes of segregants related to molybdate resistance, demonstrating the presence of segregating variation from the parental strain. Using bulk segregant analysis we mapped molybdate traits to two loci. By sequencing both the parental and evolved strain genomes we identified candidate mutations within the two regions as well as an extra copy of chromosome 1 in UMCC 2581. Combining the mapped loci with gene expression profiles of the evolved and parental strains we identified a number of candidate genes with genetic and/or gene expression changes that could underlie molybdate resistance and increased GSH levels. Our results provide insight into the genetic basis of GSH production relevant to winemaking and highlight the value of enhancing wine strains using existing variation present in wine strains.

https://www.ncbi.nlm.nih.gov/pubmed/28763437

Christian Brion, Sylvain Legrand, Jackson Peter, Claudia Caradec, David Pflieger, Jing Hou, Anne Friedrich, Bertrand Llorente, Joseph Schacherer PLoS Genet. 2017 Aug 1;13(8):e1006917. doi: 10.1371/journal.pgen.1006917. eCollection 2017 Aug. Meiotic recombination is a major factor of genome evolution, deeply characterized in only a few model species, notably the yeast Saccharomyces cerevisiae. Consequently, little is known about variations of its properties across species. In this respect, we explored the recombination landscape of Lachancea kluyveri, a protoploid yeast species that diverged from the Saccharomyces genus more than 100 million years ago and we found striking differences with S. cerevisiae. These variations include a lower recombination rate, a higher frequency of chromosomes segregating without any crossover and the absence of recombination on the chromosome arm containing the sex locus. In addition, although well conserved within the Saccharomyces clade, the S. cerevisiae recombination hotspots are not conserved over a broader evolutionary distance. Finally and strikingly, we found evidence of frequent reversal of commitment to meiosis, resulting in return to mitotic growth after allele shuffling. Identification of this major but underestimated evolutionary phenomenon illustrates the relevance of exploring non-model species.

https://www.ncbi.nlm.nih.gov/pubmed/28763437

Matthias Meurer, Veronika Chevyreva, Bram Cerulus, Michael Knop PLoS Genet. 2017 Aug 1;13(8):e1006917. doi: 10.1371/journal.pgen.1006917. eCollection 2017 Aug. Here we describe a set of tools to facilitate the use of maltose and the MAL32 promoter for regulated gene expression in yeast, alone or in combination with the GAL1 promoter. Using fluorescent protein reporters we find that under non-inducing conditions the MAL32 promoter exhibits a low basal level of expression, similar to the GAL1 promoter, and that both promoters can be induced independently of each other using the respective sugars, maltose and galactose. While their repression upon glucose addition is immediate and complete, we found that the MAL32 and GAL1 promoters each exhibit distinct induction kinetics. A set of plasmids is available to facilitate the application of the MAL32 promoter for chromosomal modifications using PCR targetting and for plasmid based gene expression.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6059479/

Tessy Korthout, Deepani W. Poramba-Liyanage, Ila van Kruijsbergen, Kitty F. Verzijlbergen, Frank P. A. van Gemert, Tibor van Welsem, Fred van Leeuwen. PLoS Biol. 2018 Jul; 16(7): e2005542. Transcription, replication, and repair involve interactions of specific genomic loci with many different proteins. How these interactions are orchestrated at any given location and under changing cellular conditions is largely unknown because systematically measuring protein–DNA interactions at a specific locus in the genome is challenging. To address this problem, we developed Epi-Decoder, a Tag-chromatin immunoprecipitation-Barcode-Sequencing (TAG-ChIP-Barcode-Seq) technology in budding yeast. Epi-Decoder is orthogonal to proteomics approaches because it does not rely on mass spectrometry (MS) but instead takes advantage of DNA sequencing. Analysis of the proteome of a transcribed locus proximal to an origin of replication revealed more than 400 interacting proteins. Moreover, replication stress induced changes in local chromatin proteome composition prior to local origin firing, affecting replication proteins as well as transcription proteins. Finally, we show that native genomic loci can be decoded by efficient construction of barcode libraries assisted by clustered regularly interspaced short palindromic repeats and CRISPR-associated protein 9 (CRISPR/Cas9). Thus, Epi-Decoder is an effective strategy to identify and quantify in an unbiased and systematic manner the proteome of an individual genomic locus by DNA sequencing.

https://www.ncbi.nlm.nih.gov/pubmed/?term=The+hidden+complexity+of+Mendelian+traits+across+yeast+natural+populations

Jing Hou, Anastasie Sigwalt, David Pflieger, Jackson Peter, Jacky de Montigny, Maitreya Dunham, Joseph Schacherer Cell Rep. 2016 Jul 26;16(4):1106-1114. doi: 10.1016/j.celrep.2016.06.048. Epub 2016 Jul 7. Mendelian traits are considered to be at the lower end of the complexity spectrum of heritable phenotypes. However, more than a century after the rediscovery of Mendel's law, the global landscape of monogenic variants, as well as their effects and inheritance patterns within natural populations, is still not well understood. Using the yeast Saccharomyces cerevisiae, we performed a species-wide survey of Mendelian traits across a large population of isolates. We generated offspring from 41 unique parental pairs and analyzed 1,105 cross/trait combinations. We found that 8.9% of the cases were Mendelian. Further tracing of causal variants revealed background-specific expressivity and modified inheritances, gradually transitioning from Mendelian to complex traits in 30% of the cases. In fact, when taking into account the natural population diversity, the hidden complexity of traits could be substantial, confounding phenotypic predictability even for simple Mendelian traits.

https://www.ncbi.nlm.nih.gov/pubmed/28751498

Jochen Kustermann, Yehui Wu, Lucia Rieger, Dirk Dedden, Tamara Phan, Paul Walther, Alexander Dünkler, Nils Johnsson J Cell Sci. 2017 Sep 15;130(18):2996-3008. doi: 10.1242/jcs.206334. Epub 2017 Jul 27. Eukaryotic cells can direct secretion to defined regions of their plasma membrane. These regions are distinguished by an elaborate architecture of proteins and lipids that are specialized to capture and fuse post-Golgi vesicles. Here, we show that the proteins Boi1p and Boi2p are important elements of this area of active exocytosis at the tip of growing yeast cells. Cells lacking Boi1p and Boi2p accumulate secretory vesicles in their buds. The essential PH domains of Boi1p and Boi2p interact with Sec1p, a protein required for SNARE complex formation and vesicle fusion. Sec1p loses its tip localization in cells depleted of Boi1p and Boi2p but overexpression of Sec1p can partially compensate for their loss. The capacity to simultaneously bind phospholipids, Sec1p, multiple subunits of the exocyst, Cdc42p and the module for generating active Cdc42p identify Boi1p and Boi2p as essential mediators between exocytosis and polar growth.

https://www.ncbi.nlm.nih.gov/pubmed/30097472

Naseeb S, Alsammar H, Burgis T, Donaldson I, Knyazev N, Knight C, Delneri D. G3 (Bethesda). 2018 Aug 30;8(9):2967-2977. doi: 10.1534/g3.118.200476. Saccharomyces sensu stricto complex consist of yeast species, which are not only important in the fermentation industry but are also model systems for genomic and ecological analysis. Here, we present the complete genome assemblies of Saccharomyces jurei, a newly discovered Saccharomyces sensu stricto species from high altitude oaks. Phylogenetic and phenotypic analysis revealed that S. jurei is more closely related to S. mikatae, than S. cerevisiae, and S. paradoxus The karyotype of S. jurei presents two reciprocal chromosomal translocations between chromosome VI/VII and I/XIII when compared to the S. cerevisiae genome. Interestingly, while the rearrangement I/XIII is unique to S. jurei, the other is in common with S. mikatae strain IFO1815, suggesting shared evolutionary history of this species after the split between S. cerevisiae and S. mikatae The number of Ty elements differed in the new species, with a higher number of Ty elements present in S. jurei than in S. cerevisiae Phenotypically, the S. jurei strain NCYC 3962 has relatively higher fitness than the other strain NCYC 3947T under most of the environmental stress conditions tested and showed remarkably increased fitness in higher concentration of acetic acid compared to the other sensu stricto species. Both strains were found to be better adapted to lower temperatures compared to S. cerevisiae.