Search results for " sequencing"
showing 10 items of 976 documents
Unveiling distribution patterns of freshwater phytoplankton by a next generation sequencing based approach.
2012
The recognition and discrimination of phytoplankton species is one of the foundations of freshwater biodiversity research and environmental monitoring. This step is frequently a bottleneck in the analytical chain from sampling to data analysis and subsequent environmental status evaluation. Here we present phytoplankton diversity data from 49 lakes including three seasonal surveys assessed by next generation sequencing (NGS) of 16S ribosomal RNA chloroplast and cyanobacterial gene amplicons and also compare part of these datasets with identification based on morphology. Direct comparison of NGS to microscopic data from three time-series showed that NGS was able to capture the seasonality in…
High-throughput sequencing of RNA silencing-associated small RNAs in olive (Olea europaea L.).
2011
14 páginas, 5 figuras, 3 tablas, S4 figuras, S2 tablas
Measuring RNA polymerase activity genome-wide with high-resolution run-on-based methods
2019
The biogenesis of RNAs is a multi-layered and highly regulated process that involves a diverse set of players acting in an orchestrated manner throughout the transcription cycle. Transcription initiation, elongation and termination factors act on RNA polymerases to modulate their movement along the DNA template in a very precise manner, more complex than previously anticipated. Genome-scale run-on-based methodologies have been developed to study in detail the position of transcriptionally-engaged RNA polymerases. Genomic run-on (GRO), and its many variants and refinements made over the years, are helping the community to address an increasing amount of scientific questions, spanning an incr…
A method for genome-wide analysis of DNA helical tension by means of psoralen-DNA photobinding
2010
The helical tension of chromosomal DNA is one of the epigenetic landmarks most difficult to examine experimentally. The occurrence of DNA crosslinks mediated by psoralen photobinding (PB) stands as the only suitable probe for assessing this problem. PB is affected by chromatin structure when is done to saturation; but it is mainly determined by DNA helical tension when it is done to very low hit conditions. Hence, we developed a method for genome-wide analysis of DNA helical tension based on PB. We adjusted in vitro PB conditions that discern DNA helical tension and applied them to Saccharomyces cerevisiae cells. We selected the in vivo cross-linked DNA sequences and identified them on DNA …
A genomic view of mRNA turnover in yeast
2011
The steady-state mRNA level is the result of two opposing processes: transcription and degradation; both of which can provide important points to regulate gene expression. In the model organism yeast Saccharomyces cerevisiae, it is now possible to determine, at the genomic level, the transcription and degradation rates, as well as the mRNA amount, using DNA chip or parallel sequencing technologies. In this way, the contribution of both rates to individual and global gene expressions can be analysed. Here we review the techniques used for the genomic evaluation of the transcription and degradation rates developed for this yeast, and we discuss the integration of the data obtained to fully an…
Pharmacological genome demethylation increases radiosensitivity of head and neck squamous carcinoma cells
2011
Aberrant inactivation of tumor suppressor genes by promoter hypermethylation has been recognized as a crucial step of tumor development and is related to aggressiveness and therapy resistance. To identify potential novel treatment strategies, we evaluated pharmacological genome demethylation for the increase of irradiation treatment effectiveness in head and neck squamous cell carcinoma (HNSCC) in this in vitro study. HNSCC cells were cultured with 2 different concentrations of 5-azacytidine (5-Aza) for 72 h, followed by a single fraction irradiation with 4 or 50 Gy, respectively. To show successful genome demethylation, the methylation status of the tumor suppressor gene hic1 (hypermethyla…
Evolution of common ragweed (Ambrosia artemisiifolia) resistance to herbicides : identification of genetic determinisms and application to molecular …
2022
Common ragweed (Ambrosia artemisiifolia L.), a particularly troublesome and allergenic weed, is mainly controlled in agricultural fields using ALS inhibitor herbicides. Recent cases of herbicide resistance have been reported in France and are jeopardising the efficacy of this mode of action. Both target site resistance (TSR, structural mutation in ALS gene) and non target site resistance (NTSR, regulatory and/or structural mutations in secondary metabolism) are involved. The fundamental aim of this work was to identify the genetic determinisms of resistance to ALS inhibitors that have evolved in common ragweed populations in France. As an applied objective, this work also aimed to prepare t…
Immunological Methods for Analysis of Recombinant Proteins
1998
We want to introduce researchers to techniques that help to solve some problems in the work of the molecular biologist. After transformation of recombinant DNA in E. coli cells many clones are usually produced, and the same situation appears if recombinant DNA expression libraries are available. Furthermore, if appropriate monoclonal or polyclonal antibodies are available, the method of immunoscreening of colonies for direct immunological detection of translational products of cloned genes can be used. Selected clones could be chosen for further studies such as determination of their primary structure by DNA sequencing and for characterization of an appropriate expressed protein.
TAF-ChIP: An ultra-low input approach for genome wide chromatin immunoprecipitation assay
2018
Chromatin immunoprecipitation (ChIP) followed by next generation sequencing is an invaluable and powerful technique to understand transcriptional regulation. However, ChIP is currently limited by the requirement of large amount of starting material. This renders studying rare cell populations very challenging, or even impossible. Here, we present a tagmentation-assisted fragmentation ChIP (TAF-ChIP) and sequencing method to generate high-quality datasets from low cell numbers. The method relies on Tn5 transposon activity to fragment the chromatin that is immunoprecipitated, thus circumventing the need for sonication or MNAse digestion to fragment. Furthermore, Tn5 adds the sequencing adapto…
The Norway spruce genome sequence and conifer genome evolution
2013
Conifers have dominated forests for more than 200 million years and are of huge ecological and economic importance. Here we present the draft assembly of the 20-gigabase genome of Norway spruce (Picea abies), the first available for any gymnosperm. The number of well-supported genes (28,354) is similar to the >100 times smaller genome of Arabidopsis thaliana, and there is no evidence of a recent whole-genome duplication in the gymnosperm lineage. Instead, the large genome size seems to result from the slow and steady accumulation of a diverse set of long-terminal repeat transposable elements, possibly owing to the lack of an efficient elimination mechanism. Comparative sequencing of Pinu…