Gene editing in porcine embryos using a combination of electroporation and transfection methods.

Autor: Takebayashi K; Bio-Innovation Research Center, Tokushima University, Tokushima, Japan.; Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima, Japan., Wittayarat M; Faculty of Veterinary Science, Prince of Songkla University, Songkhla, Thailand., Lin Q; Bio-Innovation Research Center, Tokushima University, Tokushima, Japan.; Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima, Japan., Hirata M; Bio-Innovation Research Center, Tokushima University, Tokushima, Japan.; Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima, Japan., Yoshimura N; Bio-Innovation Research Center, Tokushima University, Tokushima, Japan.; Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima, Japan., Torigoe N; Bio-Innovation Research Center, Tokushima University, Tokushima, Japan.; Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima, Japan., Nagahara M; NOSAI Yamagata Central Veterinary Clinic Center, Yamagata, Japan., Do LTK; Faculty of Veterinary Medicine, Vietnam National University of Agriculture, Hanoi, Vietnam., Tanihara F; Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima, Japan., Otoi T; Bio-Innovation Research Center, Tokushima University, Tokushima, Japan.; Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima, Japan.
Jazyk: angličtina
Zdroj: Reproduction in domestic animals = Zuchthygiene [Reprod Domest Anim] 2022 Oct; Vol. 57 (10), pp. 1136-1142. Date of Electronic Publication: 2022 Jun 22.
DOI: 10.1111/rda.14184
Abstrakt: CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats-CRISPR-associated protein 9) technology is growing rapidly and has been greatly influencing the efficiency and effectiveness of genetic modifications in different applications. One aspect of research gaining importance in the development of the CRISPR/Cas9 system is the introduction of CRISPR materials into target organisms. Although we previously demonstrated the efficacy of electroporation- and lipofection-mediated CRISPR/Cas9 gene disruption in porcine zygotes, we still believe that the efficiency of this system could be improved by combining these two methods. The present study was thus conducted to clarify the effects of a combination of electroporation and lipofection for delivering CRISPR/Cas9 components into zona pellucida (ZP)-intact and -free zygotes. The results revealed that electroporation alone significantly increased the biallelic mutation rates in the resulting blastocysts compared to lipofection alone, irrespective of the presence of ZP. None of ZP-intact zygotes treated by lipofectamine alone had any mutations, suggesting that removal of the ZP is necessary for enabling CRISPR/Cas9-based genome editing via lipofection treatment in the zygotes. Additional lipofectamine treatment after electroporation did not improve the rates of total and biallelic mutations in the resulting blastocysts derived from either ZP-intact or -free zygotes.
(© 2022 Wiley-VCH GmbH.)
Databáze: MEDLINE
Externí odkaz: