Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

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Record number 535622
Title CRISPR/cas9-mediated mutagenesis of four putative symbiosis genes of the tropical tree parasponia andersonii reveals novel phenotypes
Author(s) Zeijl, Arjan Van; Wardhani, Titis A.K.; Seifi Kalhor, Maryam; Rutten, Luuk; Bu, Fengjiao; Hartog, Marijke; Linders, Sidney; Fedorova, Elena E.; Bisseling, Ton; Kohlen, Wouter; Geurts, Rene
Source Frontiers in Plant Science 9 (2018). - ISSN 1664-462X
DOI http://dx.doi.org/10.3389/fpls.2018.00284
Department(s) Laboratory of Molecular Biology
EPS
Publication type Refereed Article in a scientific journal
Publication year 2018
Keyword(s) CRISPR/Cas9 - Nodule - Parasponia andersonii - Rhizobium - Stable transformation - Symbiosis
Abstract Parasponia represents five fast-growing tropical tree species in the Cannabaceae and is the only plant lineage besides legumes that can establish nitrogen-fixing nodules with rhizobium. Comparative analyses between legumes and Parasponia allows identification of conserved genetic networks controlling this symbiosis. However, such studies are hampered due to the absence of powerful reverse genetic tools for Parasponia. Here, we present a fast and efficient protocol for Agrobacterium tumefaciens-mediated transformation and CRISPR/Cas9 mutagenesis of Parasponia andersonii. Using this protocol, knockout mutants are obtained within 3 months. Due to efficient micro-propagation, bi-allelic mutants can be studied in the T0 generation, allowing phenotypic evaluation within 6 months after transformation. We mutated four genes – PanHK4, PanEIN2, PanNSP1, and PanNSP2 – that control cytokinin, ethylene, or strigolactone hormonal networks and that in legumes commit essential symbiotic functions. Knockout mutants in Panhk4 and Panein2 displayed developmental phenotypes, namely reduced procambium activity in Panhk4 and disturbed sex differentiation in Panein2 mutants. The symbiotic phenotypes of Panhk4 and Panein2 mutant lines differ from those in legumes. In contrast, PanNSP1 and PanNSP2 are essential for nodule formation, a phenotype similar as reported for legumes. This indicates a conserved role for these GRAS-type transcriptional regulators in rhizobium symbiosis, illustrating the value of Parasponia trees as a research model for reverse genetic studies.
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