GeneORator: An Efficient Method for the Systematic Mutagenesis of Entire Genes.

Lucy Green, Nigel S Scrutton, Andrew Currin

Abstract read

In one paragraph

Article in Methods in molecular biology (Clifton, N.J.), 2022. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.

0numbers the graph read from it

0cells of the map it votes in

0citing papers in PubMed

–field-weighted citation impact

1 · What the graph read from it

What it found

Each row is one number read from the abstract, on the scale the paper reported it, with its interval. Left of the dashed line favours the treatment, right favours the comparator. Under each row is the sentence it came from. New to these charts? A ten-minute tutorial.

The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.

2 · The registry

The trial behind it

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Neither the registry nor the abstract names a trial number. If this is a trial report, that itself is worth knowing.

3 · Its place in the literature

Who cites it

0 citing papers in PubMed.

No citing paper in PubMed yet.

4 · The record

Corrections and comments

PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.

5 · Who and what money

Authors and funding

3 authors.

Lucy GreenManchester Synthetic Biology Research Centre for Fine and Speciality Chemicals (SYNBIOCHEM), Manchester Institute of Biotechnology, School of Chemistry, Faculty of Science and Engineering, University of Manchester, Manchester, UK.

Nigel S ScruttonManchester Synthetic Biology Research Centre for Fine and Speciality Chemicals (SYNBIOCHEM), Manchester Institute of Biotechnology, School of Chemistry, Faculty of Science and Engineering, University of Manchester, Manchester, UK.

Andrew CurrinManchester Institute of Biotechnology, University of Manchester, Manchester, UK. andrew.currin@manchester.ac.uk.

Funding

Biotechnology and Biological Sciences Research Council BB/M017702/1

6 · The paper itself

Abstract

Directed evolution is a powerful tool for the rapid improvement of a target protein toward a desired fitness criteria, such as activity, specificity, or stability. In order to achieve these desired improvements, it is often beneficial to subject the entirety of the protein to mutagenesis. However, the creation of such libraries by targeted methods (i.e. site-directed mutagenesis) can be a laborious and costly task. Here we outline the GeneORator method, which uses Boolean "OR" logic to introduce specific codon mutations at multiple loci in a single reaction, thereby greatly reducing the experimental workload. The method describes library synthesis using asymmetric PCR, in which mutagenic primers are designed to create OR-type mutations at multiple sites of variation in a two-step protocol. As an example, we show how this can be utilized for controlled and economical mutagenesis of every amino acid codon in a gene.

Indexed as

Directed Molecular EvolutionProteinsCodonGene LibraryMutagenesisMutagenesis, Site-DirectedCodonProteinsAsymmetric PCRDirected evolutionDNA library assemblySynthetic biologyVariant libraries

Identifiers

PMID35727446

DOI10.1093/nar/gki591

What OpenQuestion holds

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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.