ArticleMethods in molecular biology (Clifton, N.J.)2022
Site-Directed Mutagenesis Method Mediated by Cas9.
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. Cited by 2 papers.
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.
The trial behind it
Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
Neither the registry nor the abstract names a trial number. If this is a trial report, that itself is worth knowing.
Who cites it
2 citing papers in PubMed.
- Plant G proteins: Versatile signaling hubs for synergistic improvement of agronomic traits.Journal of integrative plant biology · 2026Review
- Applications of Programmable Endonucleases in Sequence- and Ligation-Independent Seamless DNA Assembly.Biomolecules · 2023Review
Corrections and comments
PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.
Authors and funding
5 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
This study presents an in vitro CRISPR/Cas9-mediated mutagenic (ICM) system that allows rapid construction of designed mutants or site-saturation mutagenesis libraries in a PCR-independent manner. The plasmid DNA is double digested with Cas9 bearing specific single guide RNAs to remove the target nucleotides. Next, T5 exonuclease excises both 5'-ends of the linearized plasmid to generate homologous regions of approximately 15 nt. Subsequently, a short dsDNA of approximately 30-50 bp containing the desired mutation cyclizes the plasmid through base pairing and introduces the mutation into the plasmid. The gaps are repaired in Escherichia coli host cells after transformation. This method is highly efficient and accurate. Both single and multiple site-directed mutagenesis can be successfully performed, especially to large sized plasmids. This method demonstrates the great potential for creating high-quality mutant libraries in directed evolution as an alternative to PCR-based saturation mutagenesis, thus facilitating research on synthetic biology.
Indexed as
Identifiers
35727450What OpenQuestion holds
Registered trials
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.