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. 2019 Jan 14;9:1957. doi: 10.3389/fpls.2018.01957

Table 1.

Recent publications using DNA-free Genome Editing approaches.

Reference Plant species Trait GE-technique Tissue Delivery Editing efficiency Off-targets Method
Woo et al., 2015 Arabidopsis thaliana, Lactuca sativa, Nicotiana attenuata, Oryza sativa POC CRISPR/Cas9 RNPs Protoplasts PEG-fusion Calli: Monoallelic mutations: 5.7%; Biallelic mutations: 40% 0/104 (2–5 MM) Targeted deep seq.
Baek et al., 2016 Chlamydomonas reinhardtii Yield, abiotic stress CRISPR/Cas9 RNPs Single cells Electroporation Cells: Up to 0.56% 0/17 (2–4 MM) Targeted deep seq.
Malnoy et al., 2016 Malus domestica, Vitis vinifera Biotic stress CRISPR/Cas9 RNPs Protoplasts PEG-fusion Protoplasts: 0.1–6.9% n.d. n.d.
Stoddard et al., 2016 Nicotiana benthamiana POC, herbicide resistance TALEN mRNA Protoplasts PEG-fusion Protoplasts: No-UTR DNA control: 70.5%, No-UTR mRNA control: 5.8% n.d. n.d.
Shin et al., 2016 C. reinhardtii POC CRISPR/Cas9 RNPs Single cells Electroporation Up to 3 × 10-5 0/333 (1–3 MM) WGS
Svitashev et al., 2016 Zea mays Male sterility, herbicide tolerance CRISPR/Cas9 RNPs Immature embryos Biolistics Embryo: Cas9 only: 0.002–0.02 DNA delivery: 0.18–0.56 RNP delivery: 0.01–0.69 n.d. n.d.
Subburaj et al., 2016 Petunia hybrid Herbicide resistance CRISPR/Cas9 RNPs Protoplasts PEG-fusion; Protoplasts: Cas9 protein: 0.03–27.13, NR-RGEN: 5.3–34.69 n.d. n.d.
Zhang et al., 2016 Triticum aestivum Yield CRISPR/Cas9 mRNA immature embryos Biolistics No. of transgen-free frequency: 0–100% No. of homozygous transgen-free frequency: 0–36.8% 0/8 (2–4 MM) PCR- RE
Ferenczi et al., 2017 C. reinhardtii POC, Gene replacement CRISPR/Cpf1 RNPs Single cells Electroporation Colonies: 0.12–16% n.d. n.d.
Grahl et al., 2017 Non-albicans candida POC CRISPR/Cas9 RNPs Single cells Electroporation n.d. n.d. n.d.
Kim et al., 2017 Glycine max, N. attenuata Fat-synthesis; POC CRISPR/Cpf1 RNPs Protoplasts PEG-fusion Soybean: Protoplasts: LbCpf1: 0–11.7% AsCpf1: 0–1.6% Tobacco: LbCpf1 and AsCpf1: < 0.1% LbCpf1 + crRNA and AsCpf1 + crRNA: < 1% 0/23 (4–6 MM) Targeted deep seq.
Liang et al., 2017 T. aestivum Yield CRISPR/Cas9 RNPs Protoplasts; immature embryos PEG fusion, Biolistics Protoplasts: 33.4 and 21.8% Embryo cells: gw2-RNPs: 0.18–0.21% pGE-TaGW2: 0.99–1% gasr7-RNP: 45.3% 0/20 (2–5 MM) Sanger seq.
Andersson et al., 2018 Solanum tuberosum Starch syntheses CRISPR/Cas9 RNPs Protoplasts PEG-fusion Protoplasts: 1–25% n.d. n.d.
Liang et al., 2018 T. aestivum, O. sativa Yield/ POC CRISPR/Cas9/Cpf1 RNPs, TALEN-Proteins Protoplasts; immature embryos PEG fusion, Biolistics Protoplasts: 10.9–33.6% n.d. n.d.

GE-Technique, Genome editing technique; RNPs, Ribonucleoproteins; WGS, Whole Genome sequencing; POC, Proof of concept; n.d., not determined.