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Modeling human point mutation diseases in Xenopus tropicalis with a modified CRISPR/Cas9 system
Author(s) -
Shi Zhaoying,
Xin Huhu,
Tian Dandan,
Lian Jingru,
Wang Jianhui,
Liu Guanghui,
Ran Rensen,
Shi Songyuan,
Zhang Zixuan,
Shi Yu,
Deng Yi,
Hou Chunhui,
Chen Yonglong
Publication year - 2019
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.201802661r
Subject(s) - point mutation , crispr , biology , xenopus , genetics , microbiology and biotechnology , genome editing , cas9 , computational biology , mutant , gene
Precise single‐base editing mXenopus tropicalis would greatly expand the utility of this true diploid frog for modeling human genetic diseases caused by point mutations. Here, we report the efficient conversion of C‐to‐T or G‐to‐A in X. tropicalis using the rat apolipoprotein B mRNA editing enzyme catalytic subunit 1–XTEN‐clustered regularly interspaced short palindromic repeat‐associated protein 9 (Cas9) nickase‐uracil DNA glycosylase inhibitor‐nuclear localization sequence base editor [base editor 3 (BE3)]. Coinjection of guide RNA and the Cas9 mutant complex mRNA into 1‐cell stage X. tropicalis embryos caused precise C‐to‐T or G‐to‐A substitution in 14 out of 19 tested sites with efficiencies of 5–75%, which allowed for easy establishment of stable lines. Targeting the conserved T‐box 5 R237 and Tyr C28 residues in X. tropicalis with the BE3 system mimicked human Holt‐Oram syndrome and oculocutaneous albinism type 1A, respectively. Our data indicate that BE3 is an easy and efficient tool for precise base editing in X. tropicalis .—Shi, Z., Xin, H., Tian, D., Lian, J., Wang, J., Liu, G., Ran, R., Shi, S., Zhang, Z., Shi, Y., Deng, Y., Hou, C., Chen, Y. Modeling human point mutation diseases in Xenopus tropicalis with a modified CRISPR/Cas9 system. FASEB J. 33,6962–6968 (2019). www.fasebj.org