Applicability of the EFSA Opinion on site‐directed nucleases type 3 for the safety assessment of plants developed using site‐directed nucleases type 1 and 2 and oligonucleotide‐directed mutagenesis

Abstract The European Commission requested the EFSA Panel on Genetically Modified Organisms (GMO) to assess whether section 4 (hazard identification) and the conclusions of EFSA's Scientific opinion on the risk assessment of plants developed using zinc finger nuclease type 3 technique (ZFN‐3) and other site‐directed nucleases (SDN) with similar function are valid for plants developed via SDN‐1, SDN‐2 and oligonucleotide‐directed mutagenesis (ODM). In delivering this Opinion, the GMO Panel compared the hazards associated with plants produced via SDN‐1, SDN‐2 and ODM with those associated with plants obtained via both SDN‐3 and conventional breeding. Unlike for SDN‐3 methods, the application of SDN‐1, SDN‐2 and ODM approaches aims to modify genomic sequences in a way which can result in plants not containing any transgene, intragene or cisgene. Consequently, the GMO Panel concludes that those considerations which are specifically related to the presence of a transgene, intragene or cisgene included in section 4 and the conclusions of the Opinion on SDN‐3 are not relevant to plants obtained via SDN‐1, SDN‐2 or ODM as defined in this Opinion. Overall, the GMO Panel did not identify new hazards specifically linked to the genomic modification produced via SDN‐1, SDN‐2 or ODM as compared with both SDN‐3 and conventional breeding. Furthermore, the GMO Panel considers that the existing Guidance for risk assessment of food and feed from genetically modified plants and the Guidance on the environmental risk assessment of genetically modified plants are sufficient but are only partially applicable to plants generated via SDN‐1, SDN‐2 or ODM. Indeed, those guidance documents’ requirements that are linked to the presence of exogenous DNA are not relevant for the risk assessment of plants developed via SDN‐1, SDN‐2 or ODM approaches if the genome of the final product does not contain exogenous DNA.

[1]  Christoph Then,et al.  Broadening the GMO risk assessment in the EU for genome editing technologies in agriculture , 2020, Environmental Sciences Europe.

[2]  Shadma Afzal,et al.  A review of CRISPR associated genome engineering: application, advances and future prospects of genome targeting tool for crop improvement , 2020, Biotechnology Letters.

[3]  A. Alok,et al.  The present and potential future methods for delivering CRISPR/Cas9 components in plants , 2020, Journal of Genetic Engineering and Biotechnology.

[4]  Baohong Zhang CRISPR/Cas9: A Robust Genome-Editing Tool with Versatile Functions and Endless Application , 2020, International journal of molecular sciences.

[5]  Saman Majeed,et al.  Latest Developed Strategies to Minimize the Off-Target Effects in CRISPR-Cas-Mediated Genome Editing , 2020, Cells.

[6]  F. Nogué,et al.  CRISPR-induced indels and base editing using the Staphylococcus aureus Cas9 in potato , 2020, bioRxiv.

[7]  David R. Liu,et al.  Genome editing with CRISPR–Cas nucleases, base editors, transposases and prime editors , 2020, Nature Biotechnology.

[8]  Xueqin Lv,et al.  Development of a DNA double-strand break-free base editing tool in Corynebacterium glutamicum for genome editing and metabolic engineering , 2020, Metabolic engineering communications.

[9]  K. Glenn,et al.  The role of conventional plant breeding in ensuring safe levels of naturally occurring toxins in food crops , 2020 .

[10]  M. Dichgans,et al.  Detection of Deleterious On-Target Effects after HDR-Mediated CRISPR Editing. , 2020, Cell reports.

[11]  G. May,et al.  Plant Genome Editing and the Relevance of Off-Target Changes1[OPEN] , 2020, Plant Physiology.

[12]  Andreas Houben,et al.  CRISPR–Cas9-mediated induction of heritable chromosomal translocations in Arabidopsis , 2020, Nature Plants.

[13]  B. Wynne,et al.  GMO regulations and their interpretation: how EFSA’s guidance on risk assessments of GMOs is bound to fail , 2020, Environmental Sciences Europe.

[14]  G. Shao,et al.  Base Editing: The Ever Expanding Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) Tool Kit for Precise Genome Editing in Plants , 2020, Genes.

[15]  David R. Liu,et al.  Prime genome editing in rice and wheat , 2020, Nature Biotechnology.

[16]  Dipali G. Sashital,et al.  CRISPR-Cas12a has widespread off-target and dsDNA-nicking effects , 2020, The Journal of Biological Chemistry.

[17]  Christoph Then,et al.  Risk assessment of genetically engineered plants that can persist and propagate in the environment , 2020, Environmental Sciences Europe.

[18]  SpCas9-NG self-targets the sgRNA sequence in plant genome editing , 2020, Nature Plants.

[19]  R. Stupar,et al.  Integration, abundance, and transmission of mutations and transgenes in a series of CRISPR/Cas9 soybean lines , 2020, BMC Biotechnology.

[20]  Yuri B Schwartz,et al.  Pervasive head-to-tail insertions of DNA templates mask desired CRISPR-Cas9–mediated genome editing events , 2020, Science Advances.

[21]  Steven M. Solomon Genome editing in animals: why FDA regulation matters , 2020, Nature Biotechnology.

[22]  Tingting Wu,et al.  Natural variation and CRISPR/Cas9‐mediated mutation in GmPRR37 affect photoperiodic flowering and contribute to regional adaptation of soybean , 2020, Plant biotechnology journal.

[23]  Alisdair R Fernie,et al.  Metabolomics should be deployed in identification and characterization of gene-edited crops. , 2020, The Plant journal : for cell and molecular biology.

[24]  David R. Liu,et al.  Evaluation and minimization of Cas9-independent off-target DNA editing by cytosine base editors , 2020, Nature Biotechnology.

[25]  C. Collonnier,et al.  The Judgment of the CJEU of 25 July 2018 on Mutagenesis: Interpretation and Interim Legislative Proposal , 2020, Frontiers in Plant Science.

[26]  E. Kmiec,et al.  Understanding the diversity of genetic outcomes from CRISPR-Cas generated homology-directed repair , 2019, Communications Biology.

[27]  Deyue Yu,et al.  Multiplex CRISPR/Cas9‐mediated metabolic engineering increases soya bean isoflavone content and resistance to soya bean mosaic virus , 2019, Plant biotechnology journal.

[28]  S. Sriram,et al.  Genome editing in wheat microspores and haploid embryos mediated by delivery of ZFN proteins and cell‐penetrating peptide complexes , 2019, Plant biotechnology journal.

[29]  Min Kyu Kim,et al.  Retroelement Insertion in a CRISPR/Cas9 Editing Site in the Early Embryo Intensifies Genetic Mosaicism , 2019, Front. Cell Dev. Biol..

[30]  David R. Liu,et al.  Search-and-replace genome editing without double-strand breaks or donor DNA , 2019, Nature.

[31]  Mickael Malnoy,et al.  Reduced fire blight susceptibility in apple cultivars using a high‐efficiency CRISPR/Cas9‐FLP/FRT‐based gene editing system , 2019, Plant biotechnology journal.

[32]  CRISPR-Cas9-based mutagenesis frequently provokes on-target mRNA misregulation , 2019, Nature Communications.

[33]  W. Terzaghi,et al.  Knockout of two BnaMAX1 homologs by CRISPR/Cas9‐targeted mutagenesis improves plant architecture and increases yield in rapeseed (Brassica napus L.) , 2019, Plant biotechnology journal.

[34]  Baohui Liu,et al.  Perspectives on the application of genome editing technologies in crop breeding. , 2019, Molecular plant.

[35]  Hui Wei,et al.  Strategies to Increase On-Target and Reduce Off-Target Effects of the CRISPR/Cas9 System in Plants , 2019, International journal of molecular sciences.

[36]  Daniel A. Tadesse,et al.  Template plasmid integration in germline genome-edited cattle , 2019, Nature Biotechnology.

[37]  Wolfgang Huber,et al.  Biological plasticity rescues target activity in CRISPR knock outs , 2019, Nature Methods.

[38]  Jeffrey D Wolt Current risk assessment approaches for environmental and food and feed safety assessment , 2019, Transgenic Research.

[39]  Simon Sretenovic,et al.  The emerging and uncultivated potential of CRISPR technology in plant science , 2019, Nature Plants.

[40]  Stéphane Deschamps,et al.  CRISPR-Cas9 Editing in Maize: Systematic Evaluation of Off-target Activity and Its Relevance in Crop Improvement , 2019, Scientific Reports.

[41]  Yanpeng Wang,et al.  CRISPR/Cas Genome Editing and Precision Plant Breeding in Agriculture. , 2019, Annual review of plant biology.

[42]  Katharina Kawall New Possibilities on the Horizon: Genome Editing Makes the Whole Genome Accessible for Changes , 2019, Front. Plant Sci..

[43]  M. Eckerstorfer,et al.  An EU Perspective on Biosafety Considerations for Plants Developed by Genome Editing and Other New Genetic Modification Techniques (nGMs) , 2019, Front. Bioeng. Biotechnol..

[44]  Q. Gao,et al.  Cytosine, but not adenine, base editors induce genome-wide off-target mutations in rice , 2019, Science.

[45]  L. Steinmetz,et al.  Cytosine base editor generates substantial off-target single-nucleotide variants in mouse embryos , 2019, Science.

[46]  J. Kanno,et al.  Exosome-mediated horizontal gene transfer occurs in double-strand break repair during genome editing , 2019, Communications Biology.

[47]  E. Vergne,et al.  Efficient Targeted Mutagenesis in Apple and First Time Edition of Pear Using the CRISPR-Cas9 System , 2019, Front. Plant Sci..

[48]  G. Davis,et al.  Improving CRISPR-Cas9 Genome Editing Efficiency by Fusion with Chromatin-Modulating Peptides. , 2019, The CRISPR journal.

[49]  Nan Wu,et al.  CRISPR-Cas9 mediated targeted disruption of FAD2–2 microsomal omega-6 desaturase in soybean (Glycine max.L) , 2019, BMC Biotechnology.

[50]  T. Sprink,et al.  DNA-Free Genome Editing: Past, Present and Future , 2019, Front. Plant Sci..

[51]  C. Kohl,et al.  What is the available evidence for the range of applications of genome-editing as a new tool for plant trait modification and the potential occurrence of associated off-target effects: a systematic map , 2018, Environmental Evidence.

[52]  Jie Zhang,et al.  Whole genome sequencing reveals rare off‐target mutations and considerable inherent genetic or/and somaclonal variations in CRISPR/Cas9‐edited cotton plants , 2018, Plant biotechnology journal.

[53]  K. Dorn,et al.  Molecular tools enabling pennycress (Thlaspi arvense) as a model plant and oilseed cash cover crop , 2018, Plant biotechnology journal.

[54]  R. Solano,et al.  Design of a bacterial speck resistant tomato by CRISPR/Cas9‐mediated editing of SlJAZ2 , 2018, Plant biotechnology journal.

[55]  Justin P Sandoval,et al.  The complex architecture and epigenomic impact of plant T-DNA insertions , 2019, PLoS genetics.

[56]  Martin J. Aryee,et al.  Activities and specificities of CRISPR/Cas9 and Cas12a nucleases for targeted mutagenesis in maize , 2018, Plant biotechnology journal.

[57]  M. Causse,et al.  Trait discovery and editing in tomato , 2018, The Plant journal : for cell and molecular biology.

[58]  Michael J. Bernstein,et al.  Revisiting Risk Governance of GM Plants: The Need to Consider New and Emerging Gene-Editing Techniques , 2018, Front. Plant Sci..

[59]  P. Hofvander,et al.  Genome editing in potato via CRISPR-Cas9 ribonucleoprotein delivery. , 2018, Physiologia plantarum.

[60]  Caixia Gao,et al.  Applications and potential of genome editing in crop improvement , 2018, Genome Biology.

[61]  R. Eils,et al.  Cell-specific CRISPR/Cas9 activation by microRNA-dependent expression of anti-CRISPR proteins , 2018, bioRxiv.

[62]  Vladimir Nekrasov,et al.  CRISPR/Cas precision: do we need to worry about off-targeting in plants? , 2018, Plant Cell Reports.

[63]  Qian-Hao Zhu,et al.  Highly Efficient Targeted Gene Editing in Upland Cotton Using the CRISPR/Cas9 System , 2018, International journal of molecular sciences.

[64]  D. Voytas,et al.  De novo domestication of wild tomato using genome editing , 2018, Nature Biotechnology.

[65]  G. Kleter,et al.  EU court casts new plant breeding techniques into regulatory limbo , 2018, Nature Biotechnology.

[66]  Yi Zheng,et al.  Recognition of CRISPR/Cas9 off‐target sites through ensemble learning of uneven mismatch distributions , 2018, Bioinform..

[67]  Xiang Ding,et al.  CRISPR/Cas9 Assisted Multiplex Genome Editing Technique in Escherichia coli. , 2018, Biotechnology journal.

[68]  Martin J. Aryee,et al.  In vivo CRISPR editing with no detectable genome-wide off-target mutations , 2018, Nature.

[69]  A. Bradley,et al.  Repair of double-strand breaks induced by CRISPR–Cas9 leads to large deletions and complex rearrangements , 2018, Nature Biotechnology.

[70]  G. Kleter,et al.  The European Union Court's Advocate General's Opinion and new plant breeding techniques , 2018, Nature Biotechnology.

[71]  Tao Zhang,et al.  A large-scale whole-genome sequencing analysis reveals highly specific genome editing by both Cas9 and Cpf1 (Cas12a) nucleases in rice , 2018, Genome Biology.

[72]  Beate Wittbrodt,et al.  Efficient single-copy HDR by 5’ modified long dsDNA donors , 2018, bioRxiv.

[73]  W. Parrott,et al.  Novel Features and Considerations for ERA and Regulation of Crops Produced by Genome Editing , 2018, Front. Bioeng. Biotechnol..

[74]  Waseem Akhtar,et al.  Kinetics and Fidelity of the Repair of Cas9-Induced Double-Strand DNA Breaks , 2018, Molecular cell.

[75]  Yunbo Luo,et al.  CRISPR/Cas9‐mediated mutagenesis of lncRNA1459 alters tomato fruit ripening , 2018, The Plant journal : for cell and molecular biology.

[76]  L. Tian,et al.  A large-scale whole-genome sequencing analysis reveals highly specific genome editing by both Cas9 and Cpf1 nucleases in rice , 2018, bioRxiv.

[77]  Daniel F. Voytas,et al.  Low‐gluten, nontransgenic wheat engineered with CRISPR/Cas9 , 2017, Plant biotechnology journal.

[78]  Sun‐mi Lee,et al.  TALEN‐mediated targeted mutagenesis of more than 100 COMT copies/alleles in highly polyploid sugarcane improves saccharification efficiency without compromising biomass yield , 2017, Plant biotechnology journal.

[79]  Yunbo Luo,et al.  Multiplexed CRISPR/Cas9‐mediated metabolic engineering of γ‐aminobutyric acid levels in Solanum lycopersicum , 2017, Plant biotechnology journal.

[80]  A. Anand,et al.  Advancing Agrobacterium-Based Crop Transformation and Genome Modification Technology for Agricultural Biotechnology. , 2018, Current topics in microbiology and immunology.

[81]  Jeffrey D Wolt,et al.  Risk associated with off-target plant genome editing and methods for its limitation , 2017, Emerging topics in life sciences.

[82]  F. Meng,et al.  Genome editing in potato plants by agrobacterium-mediated transient expression of transcription activator-like effector nucleases , 2017, Plant Biotechnology Reports.

[83]  Deniz M. Ozata,et al.  CRISPR/Cas9-mediated genome editing induces exon skipping by alternative splicing or exon deletion , 2017, Genome Biology.

[84]  T. Cooper,et al.  Unexpected consequences: exon skipping caused by CRISPR-generated mutations , 2017, Genome Biology.

[85]  Adam Lavertu,et al.  Frameshift indels introduced by genome editing can lead to in-frame exon skipping , 2017, PloS one.

[86]  Leslie S. Edwards,et al.  Mapping the genomic landscape of CRISPR–Cas9 cleavage , 2017, Nature Methods.

[87]  C. Jung,et al.  CRISPR-Cas9 Targeted Mutagenesis Leads to Simultaneous Modification of Different Homoeologous Gene Copies in Polyploid Oilseed Rape (Brassica napus)1 , 2017, Plant Physiology.

[88]  Jian Li,et al.  Rapid generation of genetic diversity by multiplex CRISPR/Cas9 genome editing in rice , 2017, Science China Life Sciences.

[89]  Beum-Chang Kang,et al.  CRISPR/Cpf1-mediated DNA-free plant genome editing , 2017, Nature Communications.

[90]  A. Archibald,et al.  Precision engineering for PRRSV resistance in pigs: Macrophages from genome edited pigs lacking CD163 SRCR5 domain are fully resistant to both PRRSV genotypes while maintaining biological function , 2017, PLoS pathogens.

[91]  Yanpeng Wang,et al.  Efficient DNA-free genome editing of bread wheat using CRISPR/Cas9 ribonucleoprotein complexes , 2017, Nature Communications.

[92]  David R. Liu,et al.  CRISPR-Based Technologies for the Manipulation of Eukaryotic Genomes , 2017, Cell.

[93]  E. Cahoon,et al.  Significant enhancement of fatty acid composition in seeds of the allohexaploid, Camelina sativa, using CRISPR/Cas9 gene editing , 2017, Plant biotechnology journal.

[94]  E. M. DeGennaro,et al.  Multiplex gene editing by CRISPR-Cpf1 through autonomous processing of a single crRNA array , 2016, Nature Biotechnology.

[95]  Rainer Fischer,et al.  Characteristics of Genome Editing Mutations in Cereal Crops. , 2017, Trends in plant science.

[96]  David R. Liu,et al.  CRISPR-Based Technologies for the Manipulation of Eukaryotic Genomes , 2017, Cell.

[97]  M. Ward,et al.  An integrated multi-omics analysis of the NK603 Roundup-tolerant GM maize reveals metabolism disturbances caused by the transformation process , 2016, Scientific Reports.

[98]  A. Banning,et al.  Random Splicing of Several Exons Caused by a Single Base Change in the Target Exon of CRISPR/Cas9 Mediated Gene Knockout , 2016, Cells.

[99]  Joshua K Young,et al.  Genome editing in maize directed by CRISPR–Cas9 ribonucleoprotein complexes , 2016, Nature Communications.

[100]  Dipali G. Sashital,et al.  Achieving Plant CRISPR Targeting that Limits Off‐Target Effects , 2016, The plant genome.

[101]  Yi Zhang,et al.  Efficient and transgene-free genome editing in wheat through transient expression of CRISPR/Cas9 DNA or RNA , 2016, Nature Communications.

[102]  D. Voytas,et al.  Geminivirus-Mediated Genome Editing in Potato (Solanum tuberosum L.) Using Sequence-Specific Nucleases , 2016, Front. Plant Sci..

[103]  David R. Liu,et al.  Programmable editing of a target base in genomic DNA without double-stranded DNA cleavage , 2016, Nature.

[104]  G. Lin,et al.  Potential pitfalls of CRISPR/Cas9‐mediated genome editing , 2016, The FEBS journal.

[105]  N. Patron,et al.  Multi-gene engineering in plants with RNA-guided Cas9 nuclease. , 2016, Current opinion in biotechnology.

[106]  J. Mozoruk,et al.  Oligonucleotide‐directed mutagenesis for precision gene editing , 2015, Plant biotechnology journal.

[107]  Feng Zhang,et al.  Improving cold storage and processing traits in potato through targeted gene knockout. , 2016, Plant biotechnology journal.

[108]  Soon Il Kwon,et al.  DNA-free genome editing in plants with preassembled CRISPR-Cas9 ribonucleoproteins , 2015, Nature Biotechnology.

[109]  D. Voytas,et al.  Non-transgenic Plant Genome Editing Using Purified Sequence-Specific Nucleases. , 2015, Molecular plant.

[110]  S. Krimsky An Illusory Consensus behind GMO Health Assessment , 2015 .

[111]  Daniel F Voytas,et al.  Efficient Virus-Mediated Genome Editing in Plants Using the CRISPR/Cas9 System. , 2015, Molecular plant.

[112]  Robert J. Schmitz,et al.  Targeted genome modifications in soybean with CRISPR/Cas9 , 2015, BMC Biotechnology.

[113]  Fern Wickson,et al.  No scientific consensus on GMO safety , 2015, Environmental Sciences Europe.

[114]  Martin J. Aryee,et al.  GUIDE-Seq enables genome-wide profiling of off-target cleavage by CRISPR-Cas nucleases , 2014, Nature Biotechnology.

[115]  P. Beetham,et al.  Oligo-Mediated Targeted Gene Editing , 2015 .

[116]  H. Puchta,et al.  Advances in New Technology for Targeted Modification of Plant Genomes , 2015, Springer New York.

[117]  J. Doudna,et al.  The new frontier of genome engineering with CRISPR-Cas9 , 2014, Science.

[118]  F. Zhang,et al.  Improved soybean oil quality by targeted mutagenesis of the fatty acid desaturase 2 gene family. , 2014, Plant biotechnology journal.

[119]  Kang Zhang,et al.  Targeted mutagenesis in Zea mays using TALENs and the CRISPR/Cas system. , 2014, Journal of genetics and genomics = Yi chuan xue bao.

[120]  Jin-Soo Kim,et al.  Cas-OFFinder: a fast and versatile algorithm that searches for potential off-target sites of Cas9 RNA-guided endonucleases , 2014, Bioinform..

[121]  R. Tuli,et al.  RNA-Guided Genome Editing for Target Gene Mutations in Wheat , 2013, G3: Genes, Genomes, Genetics.

[122]  Eli J. Fine,et al.  DNA targeting specificity of RNA-guided Cas9 nucleases , 2013, Nature Biotechnology.

[123]  Bing Yang,et al.  Demonstration of CRISPR/Cas9/sgRNA-mediated targeted gene modification in Arabidopsis, tobacco, sorghum and rice , 2013, Nucleic acids research.

[124]  David R. Liu,et al.  High-throughput profiling of off-target DNA cleavage reveals RNA-programmed Cas9 nuclease specificity , 2013, Nature Biotechnology.

[125]  Detlef Weigel,et al.  Targeted mutagenesis in the model plant Nicotiana benthamiana using Cas9 RNA-guided endonuclease , 2013, Nature Biotechnology.

[126]  George M. Church,et al.  Multiplex and homologous recombination–mediated genome editing in Arabidopsis and Nicotiana benthamiana using guide RNA and Cas9 , 2013, Nature Biotechnology.

[127]  J. Casacuberta,et al.  Site-directed nucleases: a paradigm shift in predictable, knowledge-based plant breeding. , 2013, Trends in biotechnology.

[128]  A. Chesson,et al.  Scientific opinion addressing the safety assessment of plants developed using Zinc Finger Nuclease 3 and other site-directed Nucleases with similar function , 2012 .

[129]  J. Doudna,et al.  A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity , 2012, Science.

[130]  E. Kmiec,et al.  DNA Damage Response Pathway and Replication Fork Stress During Oligonucleotide Directed Gene Editing , 2012, Molecular therapy. Nucleic acids.

[131]  Antoine Messéan,et al.  Scientific opinion addressing the safety assessment of plants developed through cisgenesis and intragenesis , 2012 .

[132]  Dennis J. Gray,et al.  Genetic engineering technologies , 2011 .

[133]  H. Kuiper,et al.  Guidance for risk assessment of food and feed from genetically modified plants , 2011 .

[134]  Efsa Publication,et al.  EFSA Panel on Genetically Modified Organisms (GMO); Draft Scientific Opinion on the assessment of allergenicity of GM plants and microorganisms and derived food and feed , 2010 .

[135]  Jan G. Schaart,et al.  Traditional plant breeding methods , 2010 .

[136]  Antoine Messéan,et al.  Guidance on the environmental risk assessment of genetically modified plants , 2010 .

[137]  S. Krauss,et al.  Cellular responses to targeted genomic sequence modification using single-stranded oligonucleotides and zinc-finger nucleases. , 2009, DNA repair.

[138]  Allison K. Wilson,et al.  The Mutational Consequences of Plant Transformation , 2006, Journal of biomedicine & biotechnology.

[139]  P. Sharp,et al.  Oligonucleotide-directed gene repair in wheat using a transient plasmid gene repair assay system , 2006, Plant Cell Reports.

[140]  B. Linke,et al.  Detection of RNA variants transcribed from the transgene in Roundup Ready soybean , 2005 .

[141]  K. Toriyama,et al.  Chimeric RNA/DNA oligonucleotide-directed gene targeting in rice , 2004, Plant Cell Reports.

[142]  Hong-Gyu Kang,et al.  Transgene structures in T-DNA-inserted rice plants , 2003, Plant Molecular Biology.

[143]  Daniel Schubert,et al.  A comprehensive characterization of single-copy T-DNA insertions in the Arabidopsis thaliana genome , 2003, Plant Molecular Biology.

[144]  D. Somers,et al.  Complete sequence analysis of transgene loci from plants transformed via microprojectile bombardment , 2003, Plant Molecular Biology.

[145]  Marc De Loose,et al.  T-DNA Integration in Arabidopsis Chromosomes. Presence and Origin of Filler DNA Sequences1[w] , 2003, Plant Physiology.

[146]  S. Diamond,et al.  Oligonucleotide-directed single-base DNA alterations in mouse embryonic stem cells , 2003, Gene Therapy.

[147]  D. J. Peterson,et al.  Engineering herbicide-resistant maize using chimeric RNA/DNA oligonucleotides , 2000, Nature Biotechnology.

[148]  G. May,et al.  A tool for functional plant genomics: chimeric RNA/DNA oligonucleotides cause in vivo gene-specific mutations. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[149]  V. Alexeev,et al.  Stable and inheritable changes in genotype and phenotype of albino melanocytes induced by an RNA-DNA oligonucleotide , 1998, Nature Biotechnology.

[150]  M. Rice,et al.  Correction of the Mutation Responsible for Sickle Cell Anemia by an RNA-DNA Oligonucleotide , 1996, Science.