Conversion tract analysis of homology-directed genome editing using oligonucleotide donors

Yinan Kan; Eric A. Hendrickson

doi:10.1007/978-1-4939-9500-4_7

Conversion tract analysis of homology-directed genome editing using oligonucleotide donors

Yinan Kan, Eric A. Hendrickson

Molecular Biology (TMED)

Research output: Chapter in Book/Report/Conference proceeding › Chapter

1 Scopus citations

Abstract

Homology-directed genome editing is the intentional alteration of an endogenous genetic locus using information from an exogenous homology donor. A conversion tract is defined as the amount of genetic information that is converted from the homology donor to a given strand of the targeted chromosomal locus. Because of this, conversion tract analysis retrospectively not only elucidates the mechanism of homology-directed genome editing but also provides valuable insights on the conversion efficiency of every nucleotide in the homology donor. Here we describe a blue fluorescent protein-to-green fluorescent protein conversion system that can be conveniently used to measure the efficiency and analyze the lengths of conversion tracts of homology-directed genome editing using oligonucleotide donors in mammalian cells.

Original language	English (US)
Title of host publication	Methods in Molecular Biology
Publisher	Humana Press Inc.
Pages	131-144
Number of pages	14
DOIs	https://doi.org/10.1007/978-1-4939-9500-4_7
State	Published - 2019

Publication series

Name	Methods in Molecular Biology
Volume	1999
ISSN (Print)	1064-3745
ISSN (Electronic)	1940-6029

Keywords

Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) (CRISPR-Cas9)
Conversion tract
Gene conversion
Gene editing
Homology-directed repair (HDR)
Oligonucleotide

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AB - Homology-directed genome editing is the intentional alteration of an endogenous genetic locus using information from an exogenous homology donor. A conversion tract is defined as the amount of genetic information that is converted from the homology donor to a given strand of the targeted chromosomal locus. Because of this, conversion tract analysis retrospectively not only elucidates the mechanism of homology-directed genome editing but also provides valuable insights on the conversion efficiency of every nucleotide in the homology donor. Here we describe a blue fluorescent protein-to-green fluorescent protein conversion system that can be conveniently used to measure the efficiency and analyze the lengths of conversion tracts of homology-directed genome editing using oligonucleotide donors in mammalian cells.

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KW - Oligonucleotide

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