A seed resource for screening functionally redundant genes and isolation of new mutants impaired in co2 and aba responses

Felix Hauser, Paulo H.O. Ceciliato, Yi Chen Lin, Dan Dan Guo, J. D. Gregerson, Nazia Abbasi, Diana Youhanna, Jiyoung Park, Guillaume Dubeaux, Eilon Shani, Nusra Poomchongkho, Julian I. Schroeder*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

11 Scopus citations

Abstract

The identification of homologous genes with functional overlap in forward genetic screens is severely limited. Here, we report the generation of over 14 000 artificial microRNA (amiRNA)-expressing plants that enable screens of the functionally redundant gene space in Arabidopsis. A protocol was developed for isolating robust and reproducible amiRNA mutants. Examples of validation approaches and essential controls are presented for two new amiRNA mutants that exhibit genetically redundant phenotypes and circumvent double mutant lethality. In a forward genetic screen for abscisic acid (ABA)-mediated inhibition of seed germination, amiRNAs that target combinations of known redundant ABA receptor and SnRK2 kinase genes were rapidly isolated, providing a strong proof of principle for this approach. A new ABA-insensitive amiRNA line that targets three avirulence-induced gene 2(-like) genes was isolated . A thermal imaging screen for plants with impaired stomatal opening in response to low CO2 exposure led to the isolation of a new amiRNA targeting two essential proteasomal subunits, PAB1 and PAB2. The seed library of 11 000 T2 amiRNA lines (with 3000 lines in progress) generated here provides a new platform for forward genetic screens and has been made available to the Arabidopsis Biological Resource Center (ABRC). Optimized procedures for amiRNA screening and controls are described.

Original languageEnglish
Pages (from-to)641-651
Number of pages11
JournalJournal of Experimental Botany
Volume70
Issue number2
DOIs
StatePublished - 2019

Funding

FundersFunder number
National Science FoundationMCB1616236
National Institutes of HealthGM060396-ES010337
National Institute of Environmental Health SciencesP42ES010337
European Molecular Biology OrganizationALTF334-2018, 501100003043
European Research Council757683, 100010663
Conselho Nacional de Desenvolvimento Científico e Tecnológico203406/2014-1
Israel Science Foundation501100003977, 1832/14
China Scholarship Council501100004543, 201706580016
Ciência sem Fronteiras

    Keywords

    • Arabidopsis
    • Artificial microRNA
    • Forward genetic screen

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