Refined Homology-Directed Repair Methodological Approaches of Donorguide, a Chimeric RNA: DNA tracrRNA

被引：0
作者：
Gomez, Adriana Morales [1 ,4 ,5 ]
Thulung, Lucy Rai [2 ]
Daby, Camden L. [2 ]
Savage, Kaila S. [2 ]
Clark, Karl J. [3 ]
Ekker, Stephen C. [4 ,5 ]
机构：
[1] Mayo Clin, Ctr Clin & Translat Sci, Rochester, MN USA
[2] Mayo Clin, Dept Biochem & Mol Biol, Rochester, MN USA
[3] Texas A&M Univ, Dept Anim Sci, College Stn, TX USA
[4] Univ Texas Austin, Dept Pediat, Dell Med Sch, Austin, TX USA
[5] Univ Texas Austin, Ctr Rare Dis, Dell Med Sch, Austin, TX USA
来源：
GEN BIOTECHNOLOGY | 2024年 / 3卷 / 03期
关键词：
GENOMIC DNA; BASE; EFFICIENCY; CELLS;
D O I：
10.1089/genbio.2024.0016
中图分类号：
Q81 [生物工程学（生物技术）]; Q93 [微生物学];
学科分类号：
071005 ; 0836 ; 090102 ; 100705 ;
摘要：
The use of single-stranded oligodeoxynucleotide (ssODN) along with CRISPR-Cas9-mediated double-strand breaks (DSB) is one of the most commonly deployed methods for introducing genetic alterations, but this approach has notable limitations. Recognizing this, we have developed a protocol article that provides a step-by-step process of donorguide, a covalent fusion of trans-activating CRISPR RNA (tracrRNA) and ssODN. Donorguide has the potential to enhance the introduction of specific genetic alterations (insertion, deletion, and substitution) at a DSB, improving homology-directed repair methods from zebrafish in vivo to human cells in vitro. We also explored and discuss the impact of increasing the length of donorguide homology arms in zebrafish.
引用
收藏
页码：161 / 169
页数：9
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