Developing Multipurpose Nicotiana Crops for Molecular Farming using New Plant Breeding Techniques

Deliverables

Collection of functionally characterized intragenic elements
Report about Public Perception of NPBTs
A transferable universal GT toolbox for Nicotiana and other Solanaceae species
White paper on the assessment of NPBTs for future breeding practice
Communication Plan and Gender Action Plan
Final Communication Report
Initial report on Stakeholder information of NPBTs and Molecular Farming
Collection of editing enzymes with compared efficiencies and predictive model
Final Dissemination Report
Final report on stakeholder opinions and expected impacts of NPBTs
Final Report on Regulation and Biosafety updates
Initial Report on Regulation and Biosafety updates
Dissemination Plan

Development of Apollo web based genome browser for PacBio N. benthamiana genome assembly
Webtool hosting dedicated software and DNA part documentation
Collection of web based and audio-visual communication materials

PacBio sequencing and assembly of N. benthaminana genome

Stable N. benthamiana line expressing intronized Cas9

Publications

Engineering CRISPR/ Lb Cas12a for highly efficient, temperature‐tolerant plant gene editing

Author(s): Patrick Schindele, Holger Puchta
Published in: Plant Biotechnology Journal, Issue 18/5, 2020, Page(s) 1118-1120, ISSN 1467-7644
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/pbi.13275

Enhancing in planta gene targeting efficiencies in Arabidopsis using temperature‐tolerant CRISPR/ Lb Cas12a

Author(s): Laura Merker, Patrick Schindele, Teng‐Kuei Huang, Felix Wolter, Holger Puchta
Published in: Plant Biotechnology Journal, Issue 18/12, 2020, Page(s) 2382-2384, ISSN 1467-7644
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/pbi.13426

Strong and tunable anti-CRISPR/Cas activities in plants

Author(s): Calvache, Camilo; Vazquez-Vilar, Marta; Selma, Sara; Uranga, Mireia; Fernández-del-Carmen, Asun; Daròs, José-Antonio; Orzaez, Diego
Published in: Plant Biotechnology Journal, Issue 4, 2022, Page(s) 399-408, ISSN 1467-7644
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/pbi.13723

Multiple gene expression in plants using MIDAS-P, a versatile type II restriction-based modular expression vector

Author(s): Elizabeth C. Pinneh, Craig J. van Dolleweerd, Kathrin Göritzer, Pascal M. W. Drake, Julian K-C. Ma, Audrey Y-H. Teh
Published in: Biotechnology and Bioengineering, 2022, Page(s) 1660 - 1672, ISSN 0006-3592
Publisher: Wiley - V C H Verlag GmbbH & Co.
DOI: 10.1002/bit.28073

Contributions of the international plant science community to the fight against infectious diseases inhumans—part 2: Affordable drugs in edible plants forendemic and re-emerging diseases

Author(s): He, Wenshu; Baysal, Can; Lobato Gómez, María; Huang, Xin; Alvarez, Derry; Zhu, Changfu; Armario-Najera, Victoria; Blanco Perera, Amaya; Cerda Bennasser, Pedro; Saba-Mayoral, Andrea; Sobrino-Mengual, Guillermo; Vargheese, Ashwin; Abranches, Rita; Abreu, Isabel Alexandra; Balamurugan, Shanmugaraj; Bock, Ralph; Buyel, Johannes F.; da Cunha, Nicolau B.; Daniell, Henry; Faller, Roland; Folgado, Andr
Published in: Plant Biotechnology Journal, Issue 19, 2021, Page(s) 1921-1936, ISSN 1467-7644
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/pbi.13658

Plant molecular farming for the production of valuable proteins – Critical evaluation of achievements and future challenges

Author(s): Stefan Schillberg, Ricarda Finnern
Published in: Journal of Plant Physiology, Issue 258-259, 2021, Page(s) 153359, ISSN 0176-1617
Publisher: Elsevier BV
DOI: 10.1016/j.jplph.2020.153359

Metabolomics should be deployed in the identification and characterization of gene‐edited crops

Author(s): Paul D. Fraser, Asaph Aharoni, Robert D. Hall, Sanwen Huang, James J. Giovannoni, Uwe Sonnewald, Alisdair R. Fernie
Published in: The Plant Journal, Issue 102/5, 2020, Page(s) 897-902, ISSN 0960-7412
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/tpj.14679

Efficient Cas9 multiplex editing using unspaced sgRNA arrays engineering in a Potato virus X vector

Author(s): Mireia Uranga, Verónica Aragonés, Sara Selma, Marta Vázquez‐Vilar, Diego Orzáez, José‐Antonio Daròs
Published in: The Plant Journal, 2021, Page(s) 555–565, ISSN 0960-7412
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/tpj.15164

Targeted genome editing of plants and plant cells for biomanufacturing

Author(s): Buyel, Johannes F; Stöger, Eva; Bortesi, Luisa
Published in: Transgenic Research, Issue 2, 2021, Page(s) 401–426, ISSN 0962-8819
Publisher: Chapman & Hall
DOI: 10.1007/s11248-021-00236-z

New Hope for a “Cursed” Crop? Understanding Stakeholder Attitudes to Plant Molecular Farming With Modified Tobacco in Europe

Author(s): Jonathan Menary, Mario Amato, Andrés Cid Sanchez, Matthew Hobbs, Agata Pacho, Sebastian S. Fuller
Published in: Frontiers in Plant Science, Issue 11, 2020, ISSN 1664-462X
Publisher: Frontiers Media S. A.
DOI: 10.3389/fpls.2020.00791

Metabolic effects of agro-infiltration on N. benthamiana accessions

Author(s): Drapal. Margit; Enfissi, Eugenia M.A.; Fraser, Paul D.
Published in: Transgenic Research, 2021, Page(s) 303-315, ISSN 0962-8819
Publisher: Chapman & Hall
DOI: 10.1007/s11248-021-00256-9

A copper switch for inducing CRISPR/Cas9-based transcriptional activation tightly regulates gene expression in Nicotiana benthamiana

Author(s): Garcia-Perez, Elena; Diego-Martin, Borja; Quijano-Rubio, Alfredo; Moreno-Giménez, Elena; Selma, Sara; Orzaez, Diego; Vazquez-Vilar, Marta
Published in: BMC Biotechnology, Issue 22 (12), 2022, ISSN 1472-6750
Publisher: BioMed Central
DOI: 10.1186/s12896-022-00741-x

Expanding the genome-targeting scope and the site selectivity of high-precision base editors

Author(s): Junjie Tan, Fei Zhang, Daniel Karcher, Ralph Bock
Published in: Nature Communications, Issue 11/1, 2020, ISSN 2041-1723
Publisher: Nature Publishing Group
DOI: 10.1038/s41467-020-14465-z

Contributions of the international plant sciencecommunity to the fight against human infectious diseases–part 1: epidemic and pandemic diseases

Author(s): Lobato Gómez, María; Huang, Xin; Alvarez, Derry; He, Wenshu; Baysal, Can; Zhu, Changfu; Armario-Najera, Victoria; Blanco Perera, Amaya; Cerda Bennasser, Pedro; Saba-Mayoral, Andera; Sobrino-Mengual, Guillermo; Vargheese, Ashwin; Abranches, Rita; Abreu, Isabel Alexandra; Balamurugan, Shanmugaraj; Bock, Ralph; Buyel, Johannes.F.; da Cunha, Nicolau B.; Daniell, Henry; Faller, Roland; Folgado, Andr
Published in: Plant Biotechnology Journal, Issue 19, 2021, Page(s) 1901–1920, ISSN 1467-7644
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/pbi.13657

Shotguns vs Lasers: Identifying barriers and facilitators to scaling-up plant molecular farming for high-value health products

Author(s): Jonathan Menary, Matthew Hobbs, Sara Mesquita de Albuquerque, Agata Pacho, Pascal M. W. Drake, Alison Prendiville, Julian K-C. Ma, Sebastian S. Fuller
Published in: PLOS ONE, Issue 15/3, 2020, Page(s) e0229952, ISSN 1932-6203
Publisher: Public Library of Science
DOI: 10.1371/journal.pone.0229952

Pilot Production of SARS-CoV-2 Related Proteins in Plants: A Proof of Concept for Rapid Repurposing of Indoor Farms Into Biomanufacturing Facilities

Author(s): Borja Diego-Martin, Beatriz González, Marta Vazquez-Vilar, Sara Selma, Rubén Mateos-Fernández, Silvia Gianoglio, Asun Fernández-del-Carmen, Diego Orzáez
Published in: Frontiers in Plant Science, Issue 11, 2020, ISSN 1664-462X
Publisher: Frontiers Media S. A.
DOI: 10.3389/fpls.2020.612781

ABCC transporters mediate the vacuolar accumulation of crocins in saffron stigmas

Author(s): Olivia Costantina Demurtas, Rita Francisco, Gianfranco Diretto, Paola Ferrante, Sarah Frusciante, Marco Pietrella, Giuseppe Aprea, Lorenzo Borghi, Mistianne Feeney, Lorenzo Frigerio, Adriana Coricello, Giosuè Costa, Stefano Alcaro, Enrico Martinoia, Giovanni Giuliano
Published in: The Plant Cell, 2019, Page(s) tpc.00193.2019, ISSN 1040-4651
Publisher: American Society of Plant Biologists
DOI: 10.1105/tpc.19.00193

Efficient gene targeting in Nicotiana tabacum using CRISPR/SaCas9 and temperature tolerant LbCas12a

Author(s): Teng‐Kuei Huang, Brittney Armstrong, Patrick Schindele, Holger Puchta
Published in: Plant Biotechnology Journal, 2021, ISSN 1467-7644
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/pbi.13546

The chemotype core collection of genus Nicotiana

Author(s): Margit Drapal, Eugenia M. A. Enfissi, Paul D. Fraser
Published in: The Plant Journal, 2022, Page(s) 1516-1528, ISSN 0960-7412
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/tpj.15745

Engineering the N-glycosylation pathway of Nicotiana tabacum for molecular pharming using CRISPR/Cas9

Author(s): Göritzer K, Grandits M, Grünwald-Gruber C, Figl R, Mercx S, Navarre C, Ma JK-C and Teh AY-H
Published in: Frontiers in Plant Science, 2022, ISSN 1664-462X
Publisher: Frontiers Media S. A.
DOI: 10.3389/fpls.2022.1003065

Heterologous expression of Bixa orellana cleavage dioxygenase 4–3 drives crocin but not bixin biosynthesis

Author(s): Frusciante, Sarah; Demurtas, Olivia; Sulli, Maria; Mini, Paola; Aprea, Giuseppe; Diretto, Gianfranco; Karcher, Daniel; Bock, Ralph; Giuliano, Giovanni
Published in: Plant Physiology, 2021, ISSN 0032-0889
Publisher: American Society of Plant Biologists
DOI: 10.1093/plphys/kiab583

Assessment of Cas12a‐mediated gene editing efficiency in plants

Author(s): Joan Miquel Bernabé‐Orts, Iván Casas‐Rodrigo, Eugenio G. Minguet, Viola Landolfi, Victor Garcia‐Carpintero, Silvia Gianoglio, Marta Vázquez‐Vilar, Antonio Granell, Diego Orzaez
Published in: Plant Biotechnology Journal, 2019, ISSN 1467-7644
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/pbi.13113

Novel biotechnological approaches to produce biological compounds: challenges and opportunities for science communication

Author(s): Lei Pei, Markus Schmidt
Published in: Current Opinion in Biotechnology, Issue 56, 2019, Page(s) 43-47, ISSN 0958-1669
Publisher: Elsevier BV
DOI: 10.1016/j.copbio.2018.08.012

Activity‐based proteomics reveals nine target proteases for the recombinant protein‐stabilizing inhibitor Sl CYS 8 in Nicotiana benthamiana

Author(s): Philippe V. Jutras, Friederike Grosse‐Holz, Farnusch Kaschani, Markus Kaiser, Dominique Michaud, Renier A.L. Hoorn
Published in: Plant Biotechnology Journal, Issue 17, 2019, Page(s) 1670–1678, ISSN 1467-7644
Publisher: Blackwell Publishing Inc.
DOI: 10.1111/pbi.13092

Engineering of high-precision base editors for site-specific single nucleotide replacement

Author(s): Junjie Tan; Fei Zhang; Daniel Karcher; Ralph Bock
Published in: Nature Communications, Issue 1, 2019, ISSN 2041-1723
Publisher: Nature Publishing Group
DOI: 10.5281/zenodo.2586572

High-level expression of the HIV entry inhibitor griffithsin from the plastid genome and retention of biological activity in dried tobacco leaves

Author(s): Matthijs Hoelscher, Nadine Tiller, Audrey Y.-H. Teh, Guo-Zhang Wu, Julian K-C. Ma, Ralph Bock
Published in: Plant Molecular Biology, Issue 97/4-5, 2018, Page(s) 357-370, ISSN 0167-4412
Publisher: Kluwer Academic Publishers
DOI: 10.1007/s11103-018-0744-7

Construction of a fusion enzyme for astaxanthin formation and its characterisation in microbial and plant hosts: A new tool for engineering ketocarotenoids

Author(s): Marilise Nogueira, Eugenia M.A. Enfissi, Ralf Welsch, Peter Beyer, Matias D. Zurbriggen, Paul D. Fraser
Published in: Metabolic Engineering, Issue 52, 2019, Page(s) 243-252, ISSN 1096-7176
Publisher: Academic Press
DOI: 10.1016/j.ymben.2018.12.006

Critical Analysis of the Commercial Potential of Plants for the Production of Recombinant Proteins

Author(s): Stefan Schillberg, Nicole Raven, Holger Spiegel, Stefan Rasche, Matthias Buntru
Published in: Frontiers in Plant Science, Issue 10, 2019, ISSN 1664-462X
Publisher: Frontiers Media S. A.
DOI: 10.3389/fpls.2019.00720

CRISPR-Cas12a Genome Editing at the Whole-Plant Level Using Two Compatible RNA Virus Vectors

Author(s): Uranga, Mireia; Vazquez-Vilar, Marta; Orzáez, Diego; Darós, José-Antonio
Published in: The CRISPR Journal, 2021, Page(s) 761-769, ISSN 2573-1599
Publisher: Mary Ann Liebert, Inc., publishers
DOI: 10.1089/crispr.2021.0049

Production and use of encapsidated RNA mimics as positive control reagents for SARS-CoV-2 RT-qPCR diagnostics

Author(s): Peyret, Hadrien; Groppelli, Elisabetta; Clark, David; Eckersley, Nicholas; Planche, Tim; Ma, Julian; Lomonossoff, George P.
Published in: Journal of Virological Methods, 2021, ISSN 0166-0934
Publisher: Elsevier BV
DOI: 10.1016/j.jviromet.2021.114372

Identification of Transgene-Free CRISPR-Edited Plants of Rice, Tomato, and Arabidopsis by Monitoring DsRED Fluorescence in Dry Seeds

Author(s): Norma Aliaga-Franco, Cunjin Zhang, Silvia Presa, Anjil K. Srivastava, Antonio Granell, David Alabadí, Ari Sadanandom, Miguel A. Blázquez, Eugenio G. Minguet
Published in: Frontiers in Plant Science, Issue 10, 2019, ISSN 1664-462X
Publisher: Frontiers Media S. A.
DOI: 10.3389/fpls.2019.01150

The GB4.0 Platform, an All-In-One Tool for CRISPR/Cas-Based Multiplex Genome Engineering in Plants

Author(s): Vazquez-Vilar M, Garcia-Carpintero V, Selma S, Bernabé-Orts JM, Sanchez-Vicente J, Salazar-Sarasua B, Ressa A, de Paola C, Ajenjo M, Quintela JC, Fernández-del-Carmen A, Granell A and Orzáez D
Published in: Frontiers in Plant Science, 2021, ISSN 1664-462X
Publisher: Frontiers Media S. A.
DOI: 10.3389/fpls.2021.689937

Novel CRISPR/Cas applications in plants: from prime editing to chromosome engineering

Author(s): Huang, Teng-Kuei; Puchta, Holger
Published in: Transgenic Research, Issue 3, 2021, Page(s) 529–549, ISSN 0962-8819
Publisher: Chapman & Hall
DOI: 10.1007/s11248-021-00238-x

GB_SynP: A Modular dCas9-Regulated Synthetic Promoter Collection for Fine-Tuned Recombinant Gene Expression in Plants

Author(s): Moreno-Giménez, Elena; Selma, Sara; Calvache, Camilo; Orzaez, Diego
Published in: ACS Synthetic Biology, Issue 11, 2022, Page(s) 3037−3048, ISSN 2161-5063
Publisher: American Chemical Society
DOI: 10.1021/acssynbio.2c00238

A multi-omic Nicotiana benthamiana resource for fundamental research and biotechnology

Author(s): Buddhini Ranawaka, Jiyuan An, Michał T. Lorenc, Hyungtaek Jung, Maria Sulli, Giuseppe Aprea, Sally Roden, Satomi Hayashi, Tal Cooper, Zacharie LeBlanc, Victor Llaca, Diego Orzaez, Aureliano Bombarely, Julia Bally, Christopher Winefield, Giovanni Giuliano, Peter M. Waterhouse
Published in: bioRxiv, 2022, ISSN 2692-8205
Publisher: Cold Spring Harbor
DOI: 10.1101/2022.12.30.521993

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