1. First‐generation genome editing in potato using hairy root transformation
- Author
-
Jiming Jiang, Nathaniel M. Butler, and Shelley Jansky
- Subjects
0106 biological sciences ,0301 basic medicine ,Phytoene desaturase ,Solanum chacoense ,Agrobacterium ,Transgene ,Csy4 ,reagent delivery ,Plant Science ,Biology ,Plant Roots ,01 natural sciences ,03 medical and health sciences ,Transformation, Genetic ,Genome editing ,crop ,Trex2 ,plant transformation ,Gene ,Research Articles ,Solanum tuberosum ,Gene Editing ,Genetics ,targeted mutagenesis ,fungi ,Cas ,food and beverages ,Agrobacterium tumefaciens ,Plants, Genetically Modified ,biology.organism_classification ,Agrobacterium rhizogenes ,Transformation (genetics) ,030104 developmental biology ,CRISPR ,Agronomy and Crop Science ,Rhizobium ,Research Article ,germ‐line ,010606 plant biology & botany ,Biotechnology - Abstract
Summary Genome editing and cis‐gene breeding have rapidly accelerated crop improvement efforts, but their impacts are limited by the number of species capable of being genetically transformed. Many dicot species, including some vital potato relatives being used to accelerate breeding and genetics efforts, remain recalcitrant to standard Agrobacterium tumefaciens‐based transformation. Hairy root transformation using Agrobacterium rhizogenes (A. rhizogenes) provides an accelerated approach to generating transgenic material but has been limited to analysis of hairy root clones. In this study, strains of A. rhizogenes were tested in the wild diploid potato relative Solanum chacoense, which is recalcitrant to infection by Agrobacterium tumefaciens. One strain of A. rhizogenes MSU440 emerged as being capable of delivering a T‐DNA carrying the GUS marker and generating transgenic hairy root clones capable of GUS expression and regeneration to whole plants. CRISPR/Cas9 reagents targeting the potato PHYTOENE DESATURASE (StPDS) gene were expressed in hairy root clones and regenerated. We found that 64%–98% of transgenic hairy root clones expressing CRISPR/Cas9 reagents carried targeted mutations, while only 14%–30% of mutations were chimeric. The mutations were maintained in regenerated lines as stable mutations at rates averaging at 38% and were capable of germ‐line transmission to progeny. This novel approach broadens the numbers of genotypes amenable to Agrobacterium‐mediated transformation while reducing chimerism in primary events and accelerating the generation of edited materials.
- Published
- 2020
- Full Text
- View/download PDF