Hairy root transformation using Agrobacterium rhizogenes as a tool for exploring cell type-specific gene expression and function using tomato as a model

以番茄为模型,利用发根农杆菌作为工具进行毛状根转化,探索细胞类型特异性基因表达和功能

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作者:Mily Ron, Kaisa Kajala, Germain Pauluzzi, Dongxue Wang, Mauricio A Reynoso, Kristina Zumstein, Jasmine Garcha, Sonja Winte, Helen Masson, Soichi Inagaki, Fernán Federici, Neelima Sinha, Roger B Deal, Julia Bailey-Serres, Siobhan M Brady

Abstract

Agrobacterium rhizogenes (or Rhizobium rhizogenes) is able to transform plant genomes and induce the production of hairy roots. We describe the use of A. rhizogenes in tomato (Solanum spp.) to rapidly assess gene expression and function. Gene expression of reporters is indistinguishable in plants transformed by Agrobacterium tumefaciens as compared with A. rhizogenes. A root cell type- and tissue-specific promoter resource has been generated for domesticated and wild tomato (Solanum lycopersicum and Solanum pennellii, respectively) using these approaches. Imaging of tomato roots using A. rhizogenes coupled with laser scanning confocal microscopy is facilitated by the use of a membrane-tagged protein fused to a red fluorescent protein marker present in binary vectors. Tomato-optimized isolation of nuclei tagged in specific cell types and translating ribosome affinity purification binary vectors were generated and used to monitor associated messenger RNA abundance or chromatin modification. Finally, transcriptional reporters, translational reporters, and clustered regularly interspaced short palindromic repeats-associated nuclease9 genome editing demonstrate that SHORT-ROOT and SCARECROW gene function is conserved between Arabidopsis (Arabidopsis thaliana) and tomato.

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