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Sci Rep. 2015 Jul 20;5:12217. doi: 10.1038/srep12217.

Efficient CRISPR/Cas9-mediated Targeted Mutagenesis in Populus in the First Generation.

Author information

1
Key Laboratory of Eco-environments of Three Gorges Reservoir Region, Ministry of Education, Chongqing Key Laboratory of Transgenic Plant and Safety Control, Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.
2
1] Key Laboratory of Eco-environments of Three Gorges Reservoir Region, Ministry of Education, Chongqing Key Laboratory of Transgenic Plant and Safety Control, Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China [2] Key Laboratory of Adaptation and Evolution of Plateau Biota, Northwest Institute of Plateau Biology, Chinese Academy of Sciences, 810008 Xining, China.

Abstract

Recently, RNA-guided genome editing using the type II clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein (Cas) system has been applied to edit the plant genome in several herbaceous plant species. However, it remains unknown whether this system can be used for genome editing in woody plants. In this study, we describe the genome editing and targeted gene mutation in a woody species, Populus tomentosa Carr. via the CRISPR/Cas9 system. Four guide RNAs (gRNAs) were designed to target with distinct poplar genomic sites of the phytoene desaturase gene 8 (PtoPDS) which are followed by the protospacer-adjacent motif (PAM). After Agrobacterium-mediated transformation, obvious albino phenotype was observed in transgenic poplar plants. By analyzing the RNA-guided genome-editing events, 30 out of 59 PCR clones were homozygous mutants, 2 out of 59 were heterozygous mutants and the mutation efficiency at these target sites was estimated to be 51.7%. Our data demonstrate that the Cas9/sgRNA system can be exploited to precisely edit genomic sequence and effectively create knockout mutations in woody plants.

PMID:
26193631
PMCID:
PMC4507398
DOI:
10.1038/srep12217
[Indexed for MEDLINE]
Free PMC Article

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