ArticleG3 (Bethesda, Md.)2022
The MyLO CRISPR-Cas9 toolkit: a markerless yeast localization and overexpression CRISPR-Cas9 toolkit.
Article in G3 (Bethesda, Md.), 2022. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 9 papers.
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9 citing papers in PubMed, 27 citations in OpenAlex.
- Hoi1 targets BLTP2 to ER-PM contact sites to regulate lipid homeostasis.The Journal of cell biology · 2026Article
- Protein-only centromeric chromatin assembly streamlines human artificial chromosome formation.bioRxiv : the preprint server for biology · 2026Article
- Roles for the canonical polarity machinery in the de novo establishment of polarity in budding yeast spores.Molecular biology of the cell · 2025Article
- Roles for the canonical polarity machinery in thebioRxiv : the preprint server for biology · 2025Article
- Article
- Comparison of CRISPR-MAD7 and CRISPR-Cas9 for Gene Disruptions inJournal of fungi (Basel, Switzerland) · 2024Article
- Divergent directed evolution of a TetR-type repressor towards aromatic molecules.Nucleic acids research · 2023Article
- Functional expression of opioid receptors and other human GPCRs in yeast engineered to produce human sterols.Nature communications · 2022Article
- A Versatile Transcription Factor Biosensor System Responsive to Multiple Aromatic and Indole Inducers.ACS synthetic biology · 2022Article
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The genetic tractability of the yeast Saccharomyces cerevisiae has made it a key model organism for basic research and a target for metabolic engineering. To streamline the introduction of tagged genes and compartmental markers with powerful Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) - CRISPR-associated protein 9 (Cas9)-based genome editing tools, we constructed a Markerless Yeast Localization and Overexpression (MyLO) CRISPR-Cas9 toolkit with 3 components: (1) a set of optimized Streptococcus pyogenes Cas9-guide RNA expression vectors with 5 selectable markers and the option to either preclone or cotransform the gRNAs; (2) vectors for the one-step construction of integration cassettes expressing an untagged or green fluorescent protein/red fluorescent protein/hemagglutinin-tagged gene of interest at one of 3 levels, supporting localization and overexpression studies; and (3) integration cassettes containing moderately expressed green fluorescent protein- or red fluorescent protein-tagged compartmental markers for colocalization experiments. These components allow rapid, high-efficiency genomic integrations and modifications with only transient selection for the Cas9 vector, resulting in markerless transformations. To demonstrate the ease of use, we applied our complete set of compartmental markers to colabel all target subcellular compartments with green fluorescent protein and red fluorescent protein. Thus, the MyLO toolkit packages CRISPR-Cas9 technology into a flexible, optimized bundle that allows the stable genomic integration of DNA with the ease of use approaching that of transforming plasmids.
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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.