Mesoporous Silica Nanoparticle-Mediated Intracellular Cre Protein Delivery for Maize Genome Editing via <i>loxP</i> Site Excision<sup>,</sup>

Martín-Ortigosa, Susana; Peterson, David J.; Valenstein, Justin S.; Lin, Victor S.-Y.; Trewyn, Brian G.; Lyznik, L. Alexander; Wang, Kan

doi:10.1104/pp.113.233650

Cited by 200 publications

(84 citation statements)

References 55 publications

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“…In addition, the delivery of preassembled Cas9 protein-gRNA ribonucleoproteins (RNPs) (that is a DNA-free method) instead of plasmids can control Cas9 protein concentration to limit genotoxicity, can alleviate overexpression of sgRNA to reduce off-target effects, and can avoid transfer of genome-editing reagents to the next generation [79,80]. The possibility for RNP delivery to plant cells has been shown with biolistic particle bombardment where delivery of recombinase led to site-specific, heritable edits [81]. In higher plants, the traditional Agrobacterium-mediated plasmid delivery system not only can guarantee success for delivering of genome-editing reagents, but may also control transgene copy numbers to moderate concentration of sgRNA : Cas9 complexes and thus, reduce off-target activity [82].…”

Section: Methods To Minimize Off-target Effects Of Plant Genome Editingmentioning

confidence: 99%

Risk associated with off-target plant genome editing and methods for its limitation

Zhao

Wolt

2017

Emerging Topics in Life Sciences

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Assessment for potential adverse effects of plant genome editing logically focuses on the specific characteristics of the derived phenotype and its release environment. Genome-edited crops, depending on the editing objective, can be classified as either indistinguishable from crops developed through conventional plant breeding or as crops which are transgenic. Therefore, existing regulatory regimes and risk assessment procedures accommodate genome-edited crops. The ability for regulators and the public to accept a product focus in the evaluation of genome-edited crops will depend on research which clarifies the precision of the genome-editing process and evaluates unanticipated off-target edits from the process. Interpretation of genome-wide effects of genome editing should adhere to existing frameworks for comparative risk assessment where the nature and degree of effects are considered relative to a baseline of genome-wide mutations as found in crop varieties developed through conventional breeding methods. Research addressing current uncertainties regarding unintended changes from plant genome editing, and adopting procedures that clearly avoid the potential for gene drive initiation, will help to clarify anticipated public and regulatory questions regarding risk of crops derived through genome editing.

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Section: Methods To Minimize Off-target Effects Of Plant Genome Editingmentioning

confidence: 99%

Risk associated with off-target plant genome editing and methods for its limitation

Zhao

Wolt

2017

Emerging Topics in Life Sciences

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“…Targeted mutagenesis still depends on plant transformation, by which T-DNA carrying chimeric enzymes and binding motifs are integrated into the plant genome and expressed for targeted double-strand breaks [27，28, 31]. The methods independent of genetic transformation for genome editing, such as direct delivery of these reagents or transient expression of these enzymes in plant cells, will substantially simplify the process of gene editing, even in species with large genomes [32,33].…”

Section: Classic and Current Approaches To Establish Plant Gene Functionmentioning

confidence: 99%

Haploid Strategies for Functional Validation of Plant Genes

Shen

Pan

Lübberstedt

2015

Trends in Biotechnology

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Increasing knowledge of plant genome sequences requires the development of more reliable and efficient genetic approaches for genotype-phenotype validation. Functional identification of plant genes is generally achieved by a combination of creating genetic modifications and observing the according phenotype, which begins with forward-genetic methods represented by random physical and chemical mutagenesis and move towards reverse-genetic tools as targeted genome editing. A major bottleneck is time need to produce modified homozygous genotypes that can actually be used for phenotypic validation. Herein, we comprehensively address and compare available experimental approaches for functional validation of plant genes, and propose haploid strategies to reduce the time needed and cost consumed for establishing gene function. Classic and current approaches to establish plant gene functionThe classic approach for plant gene function establishment began with loss-of-function mutagenesis after treatment with mutagens such as radiation with X-rays or neutrons, or chemicals that introduce random small deletions or point mutations in plant genomes [5,8]. Chemical mutagens such as ethyl methanesulfonate (EMS) were more popular as they are less destructive, easier available, and have a higher efficiency than physical mutagens [9]. Theoretically, we can find a EMS mutation in any given gene by screening no more than 5000 plants from the mutagenized M1 generation for the model plant Arabidopsis [3,8]. Conventional mutagenesis has been widely used in forward genetic strategies that start with a phenotype of interest and address identification of genes affecting this phenotype [9][10][11].As an alternative forward-genetic tool, insertional mutagenesis, including T-DNA (Transferred DNA) and transposon tagging, facilitate the identification of genes disrupted by these elements [5]. Currently, T-DNA-tagged lines have been generated in large numbers, becoming a popular resource for plant gene function [12,13].Superior to T-DNA, mobilizable transponsons can provide a verification about mutational effects of insertions when they are remobilized from the insertion site to recover a potential phenotype [14]. Insertional mutagenesis is less practicable for species, for which a systematic transformation platform has not been established [15].As the number of characterized plant genes increases, reverse-genetic methodologies play an increasingly important role in gene function validation [16]. Targeting Induced Local Lesions in Genomes (TILLING) is the first reverse-genetic tool, in which chemical or physical mutagenesis is followed by a high-throughput screening for point mutations [3,4,16]. TILLING is practicable for plant species with large-sized genomes and without transformation system because it is not different from traditional chemical or physical mutagenesis in creating mutations [12].Changes of gene expression levels may result in modified phenotypes, which can be another powerful approach for elucidating gene function [5...

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“…Later in 2014, the same team explored direct delivery of Cre recombinase protein into maize cells using MSN carriers [61]. This study was of particular interest for genome editing since Cre recombinase can avoid DNA (transgene) integration into the genome and generate precise modifications without creating transgenic plants.…”

Section: Biolistic Methodsmentioning

confidence: 99%

Controlled release and intracellular protein delivery from mesoporous silica nanoparticles

Deodhar

Adams

Trewyn

2016

Biotechnology Journal

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Protein therapeutics are promising candidates for disease treatment due to their high specificity and minimal adverse side effects; however, targeted protein delivery to specific sites has proven challenging. Mesoporous silica nanoparticles (MSN) have demonstrated to be ideal candidates for this application, given their high loading capacity, biocompatibility, and ability to protect host molecules from degradation. These materials exhibit tunable pore sizes, shapes and volumes, and surfaces which can be easily functionalized. This serves to control the movement of molecules in and out of the pores, thus entrapping guest molecules until a specific stimulus triggers release. In this review, we will cover the benefits of using MSN as protein therapeutic carriers, demonstrating that there is great diversity in the ways MSN can be used to service proteins. Methods for controlling the physical dimensions of pores via synthetic conditions, applications of therapeutic protein loaded MSN materials in cancer therapies, delivering protein loaded MSN materials to plant cells using biolistic methods, and common stimuli‐responsive functionalities will be discussed. New and exciting strategies for controlled release and manipulation of proteins are also covered in this review. While research in this area has advanced substantially, we conclude this review with future challenges to be tackled by the scientific community.

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Mesoporous Silica Nanoparticle-Mediated Intracellular Cre Protein Delivery for Maize Genome Editing via loxP Site Excision^,

Cited by 200 publications

References 55 publications

Risk associated with off-target plant genome editing and methods for its limitation

Risk associated with off-target plant genome editing and methods for its limitation

Haploid Strategies for Functional Validation of Plant Genes

Controlled release and intracellular protein delivery from mesoporous silica nanoparticles

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Mesoporous Silica Nanoparticle-Mediated Intracellular Cre Protein Delivery for Maize Genome Editing via loxP Site Excision,

Cited by 200 publications

References 55 publications

Risk associated with off-target plant genome editing and methods for its limitation

Risk associated with off-target plant genome editing and methods for its limitation

Haploid Strategies for Functional Validation of Plant Genes

Controlled release and intracellular protein delivery from mesoporous silica nanoparticles

Contact Info

Product

Resources

About

Mesoporous Silica Nanoparticle-Mediated Intracellular Cre Protein Delivery for Maize Genome Editing via loxP Site Excision^,