PINE-TREE enables highly efficient genetic modification of human cell lines

Carlye Frisch; William W. Kostes; Brooke Galyon; Brycelyn Whitman; Stefan J. Tekel; Kylie Standage-Beier; Gayathri Srinivasan; Xiao Wang; David A. Brafman

doi:10.1016/j.omtn.2023.07.007

PINE-TREE enables highly efficient genetic modification of human cell lines

Carlye Frisch, William W. Kostes, Brooke Galyon, Brycelyn Whitman, Stefan J. Tekel, Kylie Standage-Beier, Gayathri Srinivasan, Xiao Wang, David A. Brafman

Engineering, Ira A. Fulton Schools of (IAFSE)

Research output: Contribution to journal › Article › peer-review

2 Scopus citations

Abstract

Prime editing technologies enable precise genome editing without the caveats of CRISPR nuclease-based methods. Nonetheless, current approaches to identify and isolate prime-edited cell populations are inefficient. Here, we established a fluorescence-based system, prime-induced nucleotide engineering using a transient reporter for editing enrichment (PINE-TREE), for real-time enrichment of prime-edited cell populations. We demonstrated the broad utility of PINE-TREE for highly efficient introduction of substitutions, insertions, and deletions at various genomic loci. Finally, we employ PINE-TREE to rapidly and efficiently generate clonal isogenic human pluripotent stem cell lines, a cell type recalcitrant to genome editing.

Original language	English (US)
Pages (from-to)	483-492
Number of pages	10
Journal	Molecular Therapy - Nucleic Acids
Volume	33
DOIs	https://doi.org/10.1016/j.omtn.2023.07.007
State	Published - Sep 12 2023

Keywords

CRISPR
MT: RNA/DNA
genome modification
human pluripotent stem cells
prime editing

ASJC Scopus subject areas

Molecular Medicine
Drug Discovery

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10.1016/j.omtn.2023.07.007

Cite this

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title = "PINE-TREE enables highly efficient genetic modification of human cell lines",

abstract = "Prime editing technologies enable precise genome editing without the caveats of CRISPR nuclease-based methods. Nonetheless, current approaches to identify and isolate prime-edited cell populations are inefficient. Here, we established a fluorescence-based system, prime-induced nucleotide engineering using a transient reporter for editing enrichment (PINE-TREE), for real-time enrichment of prime-edited cell populations. We demonstrated the broad utility of PINE-TREE for highly efficient introduction of substitutions, insertions, and deletions at various genomic loci. Finally, we employ PINE-TREE to rapidly and efficiently generate clonal isogenic human pluripotent stem cell lines, a cell type recalcitrant to genome editing.",

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doi = "10.1016/j.omtn.2023.07.007",

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T1 - PINE-TREE enables highly efficient genetic modification of human cell lines

AU - Frisch, Carlye

AU - Kostes, William W.

AU - Galyon, Brooke

AU - Whitman, Brycelyn

AU - Tekel, Stefan J.

AU - Standage-Beier, Kylie

AU - Srinivasan, Gayathri

AU - Wang, Xiao

AU - Brafman, David A.

PY - 2023/9/12

Y1 - 2023/9/12

N2 - Prime editing technologies enable precise genome editing without the caveats of CRISPR nuclease-based methods. Nonetheless, current approaches to identify and isolate prime-edited cell populations are inefficient. Here, we established a fluorescence-based system, prime-induced nucleotide engineering using a transient reporter for editing enrichment (PINE-TREE), for real-time enrichment of prime-edited cell populations. We demonstrated the broad utility of PINE-TREE for highly efficient introduction of substitutions, insertions, and deletions at various genomic loci. Finally, we employ PINE-TREE to rapidly and efficiently generate clonal isogenic human pluripotent stem cell lines, a cell type recalcitrant to genome editing.

AB - Prime editing technologies enable precise genome editing without the caveats of CRISPR nuclease-based methods. Nonetheless, current approaches to identify and isolate prime-edited cell populations are inefficient. Here, we established a fluorescence-based system, prime-induced nucleotide engineering using a transient reporter for editing enrichment (PINE-TREE), for real-time enrichment of prime-edited cell populations. We demonstrated the broad utility of PINE-TREE for highly efficient introduction of substitutions, insertions, and deletions at various genomic loci. Finally, we employ PINE-TREE to rapidly and efficiently generate clonal isogenic human pluripotent stem cell lines, a cell type recalcitrant to genome editing.

KW - CRISPR

KW - MT: RNA/DNA

KW - genome modification

KW - human pluripotent stem cells

KW - prime editing

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DO - 10.1016/j.omtn.2023.07.007

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EP - 492

JO - Molecular Therapy - Nucleic Acids

JF - Molecular Therapy - Nucleic Acids

ER -

PINE-TREE enables highly efficient genetic modification of human cell lines

Abstract

Keywords

ASJC Scopus subject areas

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