Application of protoplast technology to CRISPR/Cas9 mutagenesis: from single-cell mutation detection to mutant plant regeneration

doi:10.1111/pbi.12870

KIZ OpenIR > 基因起源组（王文）

	Application of protoplast technology to CRISPR/Cas9 mutagenesis: from single-cell mutation detection to mutant plant regeneration
	Choun-Sea Lin 1; Chen-Tran Hsu 1; Ling-Hung Yang 1; Lan-Ying Lee 2; Jin-Yuan Fu 1; Qiao-Wei Cheng 1; Fu-Hui Wu 1; Han C.-W. Hsiao 3; Yesheng Zhang 4; Ru Zhang 4; Wan-Jung Chang 1; Chen-Ting Yu 1; Wen Wang 4; Li-Jen Liao 5; Stanton B. Gelvin 2; Ming-Che Shih 1
	2018
发表期刊	Plant Biotechnology Journal
期号	16 页码:1295–1310
摘要	Plant protoplasts are useful for assessing the efficiency of clustered regularly interspaced shortpalindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) mutagenesis. We improved theprocess of protoplast isolation and transfection of several plant species. We also developed amethod to isolate and regenerate single mutagenized Nicotianna tabacum protoplasts intomature plants. Following transfection of protoplasts with constructs encoding Cas9 and sgRNAs,target gene DNA could be amplified for further analysis to determine mutagenesis efficiency. Weinvestigated N. tabacum protoplasts and derived regenerated plants for targeted mutagenesis ofthe phytoene desaturase (NtPDS) gene. Genotyping of albino regenerants indicated that all fourNtPDS alleles were mutated in amphidiploid tobacco, and no Cas9 DNA could be detected inmost regenerated plants.
关键词	Protoplast Isolation Crispr/cas9 Protoplast Regeneration Single-cell Analysis
DOI	10.1111/pbi.12870
语种	英语
引用统计
文献类型	期刊论文
条目标识符	http://ir.kiz.ac.cn/handle/152453/12295
专题	基因起源组（王文）
通讯作者	Choun-Sea Lin; Stanton B. Gelvin; Ming-Che Shih
作者单位	1.Agricultural Biotechnology Research Center, Academia Sinica, Taipei, Taiwan 2.Department of Biological Sciences, Purdue University, West Lafayette, IN, USA 3.Department of Bioinformatics and Medical Engineering, Asia University, Taichung City, Taiwan 4.State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China 5.Institute of Life Science, National Kaohsiung Normal University, Kaohsiung, Taiwan
推荐引用方式 GB/T 7714	Choun-Sea Lin,Chen-Tran Hsu,Ling-Hung Yang,et al. Application of protoplast technology to CRISPR/Cas9 mutagenesis: from single-cell mutation detection to mutant plant regeneration[J]. Plant Biotechnology Journal,2018(16):1295–1310.
APA	Choun-Sea Lin.,Chen-Tran Hsu.,Ling-Hung Yang.,Lan-Ying Lee.,Jin-Yuan Fu.,...&Ming-Che Shih.(2018).Application of protoplast technology to CRISPR/Cas9 mutagenesis: from single-cell mutation detection to mutant plant regeneration.Plant Biotechnology Journal(16),1295–1310.
MLA	Choun-Sea Lin,et al."Application of protoplast technology to CRISPR/Cas9 mutagenesis: from single-cell mutation detection to mutant plant regeneration".Plant Biotechnology Journal .16(2018):1295–1310.

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