Overproduction of the EcoRII endonuclease and methylase by Escherichia coli strains carrying recombinant plasmids constructed in vitro

Kosykh, V.G.; Solonin, Alexander S.; Buryanov, Yа. I.; Bayev, A.A.

doi:10.1016/0005-2787(81)90072-1

Cited by 13 publications

(3 citation statements)

References 10 publications

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“…This allowed many R–M systems to be cloned in one step, on DNA fragments that contained both genes. Among these were HhaII (173,174), EcoRII (175,176), EcoRI (164,177), PstI (178,179), PaeR7I (180–182), EcoRV (183,184), PvuII (185) and BsuRI (186). Some of these systems occurred on plasmids and were isolated by simple sub-cloning.…”

Section: Introductionmentioning

confidence: 99%

Type II restriction endonucleases—a historical perspective and more

2014

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This article continues the series of Surveys and Summaries on restriction endonucleases (REases) begun this year in Nucleic Acids Research. Here we discuss ‘Type II’ REases, the kind used for DNA analysis and cloning. We focus on their biochemistry: what they are, what they do, and how they do it. Type II REases are produced by prokaryotes to combat bacteriophages. With extreme accuracy, each recognizes a particular sequence in double-stranded DNA and cleaves at a fixed position within or nearby. The discoveries of these enzymes in the 1970s, and of the uses to which they could be put, have since impacted every corner of the life sciences. They became the enabling tools of molecular biology, genetics and biotechnology, and made analysis at the most fundamental levels routine. Hundreds of different REases have been discovered and are available commercially. Their genes have been cloned, sequenced and overexpressed. Most have been characterized to some extent, but few have been studied in depth. Here, we describe the original discoveries in this field, and the properties of the first Type II REases investigated. We discuss the mechanisms of sequence recognition and catalysis, and the varied oligomeric modes in which Type II REases act. We describe the surprising heterogeneity revealed by comparisons of their sequences and structures.

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Section: Introductionmentioning

confidence: 99%

Type II restriction endonucleases—a historical perspective and more

2014

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“…Due to the fact that the question of both the potential toxicity of chemical expression inducers and the toxicity of the final sought products is still open, the previously described variants of the initiation of biosynthesis by means of an inductor or a temperature shift are widely used. In this case, the sequence of the target gene is placed under the control of effective phage promoters [11][12][13]. To increase the solubility, expression level, and effective purification by affinity chromatography, short amino acid marker sequences called "fusion tags" are attached to the Nor C-termini of recombinant proteins [14].…”

Section: Discussionmentioning

confidence: 99%

Monoclonal Antibody HlyIIC‑15 to C-End Domain HlyII B. cereus Interacts with the Trombin Recognition Site

et al. 2020

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Among the panel of monoclonal antibodies to the recombinant protein HlyIICTD Bacillus cereus an antibody was found capable of forming an immune complex with a thrombin recognition region, the amino acid sequence of which is located inside the recombinant HlyIICTD. Localization of the epitope was carried out using peptide phage display methods, as well as enzyme immunoassay and immunoblotting for interaction with recombinant proteins, either containing or not containing individual components HlyIICTD. The identified epitope is located in the region of the thrombin site and retains the ability to interact with the antibody after the proteolyotic attack of the protein by thrombin.

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“…Cloning of the methylase EcoRII nuclear localization signal (M•EcoRII-NLS) fusion protein was performed using standard recombinant DNA techniques. Purification of this fusion protein was carried out using a modification of the procedure described previously (11,12). The procedure is representative of those employed in this work and can be summarized as follows.…”

Section: Enzymes and Chemicalsmentioning

confidence: 99%

Mobility-shift analysis with microfluidics chips

et al. 2003

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Electrophoretic mobility shift analysis (EMSA) is a well-characterized and widely used technique for the analysis of protein-DNA interaction and the analysis of transcription factor combinatorics. Currently implemented EMSA generally involves the time-consuming use of radiolabeled DNA and polyacrylamide gel electrophoresis. We are studying the bionanoscience of self-assembling supramolecular protein-nucleic nanostructures. We have undertaken these studies because they promise to enhance our understanding of assemblies formed during prebiotic evolution, provide tools for analysis of biological processes like DNA recombination, and may lead to the development of nanoscale biosensors designed for site-specific molecular targeting. During the course of that work, we noted that EMSA of these complex structures could be effectively implemented with microfluidics chips designed for the separation of DNA fragments. In this report we compare the two techniques and demonstrate that the microfluidics system is also capable of resolving complex mixtures produced by decorating DNA recombination intermediates with mixtures of DNA binding proteins. Moreover, the microfluidics chip system improves EMSA by permitting analysis with smaller samples, avoiding the use of radiolabeling, and reducing the time involved to a matter of minutes.

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Overproduction of the EcoRII endonuclease and methylase by Escherichia coli strains carrying recombinant plasmids constructed in vitro

Cited by 13 publications

References 10 publications

Type II restriction endonucleases—a historical perspective and more

Type II restriction endonucleases—a historical perspective and more

Monoclonal Antibody HlyIIC‑15 to C-End Domain HlyII B. cereus Interacts with the Trombin Recognition Site

Mobility-shift analysis with microfluidics chips

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