Plant Biosystems Design Research Roadmap 1.0

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Abstract

Human life intimately depends on plants for food, biomaterials, health, energy, and a sustainable environment. Various plants have been genetically improved mostly through breeding, along with limited modification via genetic engineering, yet they are still not able to meet the ever-increasing needs, in terms of both quantity and quality, resulting from the rapid increase in world population and expected standards of living. A step change that may address these challenges would be to expand the potential of plants using biosystems design approaches. This represents a shift in plant science research from relatively simple trial-and-error approaches to innovative strategies based on predictive models of biological systems. Plant biosystems design seeks to accelerate plant genetic improvement using genome editing and genetic circuit engineering or create novel plant systems through de novo synthesis of plant genomes. From this perspective, we present a comprehensive roadmap of plant biosystems design covering theories, principles, and technical methods, along with potential applications in basic and applied plant biology research. We highlight current challenges, future opportunities, and research priorities, along with a framework for international collaboration, towards rapid advancement of this emerging interdisciplinary area of research. Finally, we discuss the importance of social responsibility in utilizing plant biosystems design and suggest strategies for improving public perception, trust, and acceptance.

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Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems provide bacteria and archaea with adaptive immunity against viruses and plasmids by using CRISPR RNAs (crRNAs) to guide the silencing of invading nucleic acids. We show here that in a subset of these systems, the mature crRNA that is base-paired to trans-activating crRNA (tracrRNA) forms a two-RNA structure that directs the CRISPR-associated protein Cas9 to introduce double-stranded (ds) breaks in target DNA. At sites complementary to the crRNA-guide sequence, the Cas9 HNH nuclease domain cleaves the complementary strand, whereas the Cas9 RuvC-like domain cleaves the noncomplementary strand. The dual-tracrRNA:crRNA, when engineered as a single RNA chimera, also directs sequence-specific Cas9 dsDNA cleavage. Our study reveals a family of endonucleases that use dual-RNAs for site-specific DNA cleavage and highlights the potential to exploit the system for RNA-programmable genome editing.

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Daniel Gibson, Lei Young, Ray-Yuan Chuang … (2009)

We describe an isothermal, single-reaction method for assembling multiple overlapping DNA molecules by the concerted action of a 5' exonuclease, a DNA polymerase and a DNA ligase. First we recessed DNA fragments, yielding single-stranded DNA overhangs that specifically annealed, and then covalently joined them. This assembly method can be used to seamlessly construct synthetic and natural genes, genetic pathways and entire genomes, and could be a useful molecular engineering tool.

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Search-and-replace genome editing without double-strand breaks or donor DNA

Andrew Anzalone, Peyton B. Randolph, Jessie Davis … (2019)

Summary Most genetic variants that contribute to disease 1 are challenging to correct efficiently and without excess byproducts 2–5 . Here we describe prime editing, a versatile and precise genome editing method that directly writes new genetic information into a specified DNA site using a catalytically impaired Cas9 fused to an engineered reverse transcriptase, programmed with a prime editing guide RNA (pegRNA) that both specifies the target site and encodes the desired edit. We performed >175 edits in human cells including targeted insertions, deletions, and all 12 types of point mutations without requiring double-strand breaks or donor DNA templates. We applied prime editing in human cells to correct efficiently and with few byproducts the primary genetic causes of sickle cell disease (requiring a transversion in HBB) and Tay-Sachs disease (requiring a deletion in HEXA), to install a protective transversion in PRNP, and to precisely insert various tags and epitopes into target loci. Four human cell lines and primary post-mitotic mouse cortical neurons support prime editing with varying efficiencies. Prime editing shows higher or similar efficiency and fewer byproducts than homology-directed repair, complementary strengths and weaknesses compared to base editing, and much lower off-target editing than Cas9 nuclease at known Cas9 off-target sites. Prime editing substantially expands the scope and capabilities of genome editing, and in principle can correct up to 89% of known genetic variants associated with human diseases.

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Author and article information

Contributors

Xiaohan Yang:

ORCID: https://orcid.org/0000-0001-5207-4210

Journal

Journal ID (nlm-ta): Biodes Res

Journal ID (iso-abbrev): Biodes Res

Journal ID (publisher-id): BDR

Title: Biodesign Research

Publisher: AAAS

ISSN (Electronic): 2693-1257

Publication date (Electronic, pub): 5 December 2020

Publication date (Electronic, collection): 2020

Volume: 2020

Electronic Location Identifier: 8051764

Affiliations

¹Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831, USA

²The Center for Bioenergy Innovation, Oak Ridge National Laboratory, Oak Ridge, TN 37831, USA

³Department of Biology, Colorado State University, Fort Collins, CO 80523, USA

⁴Department of Plant Biology, University of California, Davis, Davis, CA, USA

⁵Feedstocks Division, Joint BioEnergy Institute, Emeryville, CA, USA

⁶Department of Biodesign, Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA

⁷Department of Chemical and Biomolecular Engineering, University of Tennessee, Knoxville, TN 37996, USA

⁸SynthSys and Institute of Molecular Plant Sciences, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK

⁹Department of Biochemistry, Genetics and Microbiology, Forestry and Agricultural Biotechnology Institute (FABI), University of Pretoria, Pretoria 0002, South Africa

¹⁰Department of Food Science, University of Copenhagen, Rolighedsvej 26, DK-1858, Frederiksberg, Copenhagen, Denmark

¹¹State Key Laboratory of Subtropical Silviculture, School of Forestry and Biotechnology, Zhejiang A&F University, Hangzhou, Zhejiang 311300, China

¹²State Key Laboratory of Tree Genetics and Breeding, Research Institute of Subtropical Forestry, Chinese Academy of Forestry, Hangzhou, Zhejiang 311400, China

¹³Department of Plant Pathology and Environmental Microbiology and the Huck Institute of the Life Sciences, The Pennsylvania State University, University Park, PA 16802, USA

¹⁴Department of Genetics, Cell Biology and Development, Center for Precision Plant Genomics and Center for Genome Engineering, University of Minnesota, Saint Paul, MN 55108, USA

¹⁵Department of Plant Science and Landscape Architecture, University of Connecticut, Storrs, CT 06269, USA

¹⁶Division of Plant Sciences, Bond Life Sciences Center, University of Missouri, Columbia, MO, USA

¹⁷Donald Danforth Plant Science Center, St. Louis, MO, USA

¹⁸Environmental Sciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831, USA