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The ability to introduce fluorophores selectively into proteins provides a powerful tool to study protein structure, dynamics, localization, and biomolecular interactions both in vitro and in vivo. Here, we report a strategy for the selective and efficient biosynthetic incorporation of a low-molecular-weight fluorophore into proteins at defined sites. The(More)
Next-generation sequencing has still not reached its full potential due to the technical inability of effectively targeting desired genomic regions of interest. Once available, methods adressing this bottleneck will dramatically reduce cost and enable the efficient analysis of complex samples. Recently, a number of possible approaches for genomic-scale(More)
We report a flexible method for selective capture of sequence fragments from complex, eukaryotic genome libraries for next-generation sequencing based on hybridization to DNA microarrays. Using microfluidic array architecture and integrated hardware, the process is amenable to complete automation and does not introduce amplification steps into the standard(More)
The lack of efficient high-throughput methods for enrichment of specific sequences from genomic DNA represents a key bottleneck in exploiting the enormous potential of next-generation sequencers. Such methods would allow for a systematic and targeted analysis of relevant genomic regions. Recent studies reported sequence enrichment using a hybridization step(More)
The fidelity of DNA polymerase activity is of central importance for numerous biotechnological applications.r1.2 1 The imperfect fidelity of DNA polymerases under the unnatural conditions of several techniques such as those of the polymerase chain reaction (PCR) either restricts the application of these enzymes or demands their tedious optimization. Thus, a(More)
A large number of amino acids other than the canonical amino acids can now be easily incorporated in vivo into proteins at genetically encoded positions. The technology requires an orthogonal tRNA/aminoacyl-tRNA synthetase pair specific for the unnatural amino acid that is added to the media while a TAG amber or frame shift codon specifies the incorporation(More)
Here, we report a generally applicable PEGylation methodology based on the site-specific incorporation of para-azidophenylalanine into proteins in yeast. The azido group was used in a mild [3+2] cycloaddition reaction with an alkyne derivatized PEG reagent to afford selectively PEGylated protein. This strategy should be useful for the generation of(More)
We genetically encoded the photocaged amino acid 4,5-dimethoxy-2-nitrobenzylserine (DMNB-Ser) in Saccharomyces cerevisiae in response to the amber nonsense codon TAG. This amino acid was converted to serine in living cells by irradiation with relatively low-energy blue light and was used to noninvasively photoactivate phosphorylation of the transcription(More)
Recent years have seen tremendous progress in next generation sequencing technologies, allowing genomic sequencing in a highly cost-effective manner. However, sample preparation for these sequencers remains a bottleneck as the human genome is too complex to be routinely resequenced. We present here an in-depth study of HybSelect, a method that can(More)
We report the genetic encoding of a noncanonical, spin-labeled amino acid in Escherichia coli. This enables the intracellular biosynthesis of spin-labeled proteins and obviates the need for any chemical labeling step usually required for protein electron paramagnetic resonance (EPR) studies. The amino acid can be introduced at multiple, user-defined sites(More)