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We describe a DNA sequencing technology in which a commonly available, inexpensive epifluorescence microscope is converted to rapid nonelectrophoretic DNA sequencing automation. We apply this technology to resequence an evolved strain of Escherichia coli at less than one error per million consensus bases. A cell-free, mate-paired library provided single DNA(More)
Genome sequencing currently requires DNA from pools of numerous nearly identical cells (clones), leaving the genome sequences of many difficult-to-culture microorganisms unattainable. We report a sequencing strategy that eliminates culturing of microorganisms by using real-time isothermal amplification to form polymerase clones (plones) from the DNA of(More)
We present genome engineering technologies that are capable of fundamentally reengineering genomes from the nucleotide to the megabase scale. We used multiplex automated genome engineering (MAGE) to site-specifically replace all 314 TAG stop codons with synonymous TAA codons in parallel across 32 Escherichia coli strains. This approach allowed us to measure(More)
We perform a genome-wide analysis of the transition between transcriptional initiation and elongation in Escherichia coli by determining the association of core RNA polymerase (RNAP) and the promoter-recognition factor sigma70 with respect to RNA transcripts. We identify 1286 sigma70-associated promoters, including many internal to known operons, and(More)
Escherichia coli MelR protein is a transcription activator that is essential for melibiose-dependent expression of the melAB genes. We have used chromatin immunoprecipitation to study the binding of MelR and RNA polymerase to the melAB promoter in vivo. Our results show that MelR is associated with promoter DNA, both in the absence and presence of the(More)
7 Corresponding authors 2 The genomic program for development depends on the precise coordination of gene activity in space and time 1. It is widely assumed that the key rate-limiting step in gene activation is the recruitment of RNA polymerase II (Pol II) to the core promoter 2. Although there are well-documented examples where Pol II is recruited to a(More)
Untreated (–) and ExoSAP-IT treated (+) PCR products were analyzed by gel electrophoresis. A variety of PCR products of different lengths may be treated with ExoSAP-IT, with no sample loss. 125 bp 455 bp 1.55 kb 4.6 kb HES-1 numb NRAGE numb M – + – + – + – + Stop losing time and samples. ExoSAP-IT ® offers quick, one step PCR clean-up with 100% recovery of(More)
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