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Development of a sequence-indexed collection of transposon tagged maize mutants as a novel public resource and its application in root developmental biology

Subject Area Plant Breeding and Plant Pathology
Term from 2019 to 2024
Project identifier Deutsche Forschungsgemeinschaft (DFG) - Project number 415165461
 
Final Report Year 2024

Final Report Abstract

Genome-wide insertional mutagenesis is a tool to generate loss-of-function mutations for virtually all genes in a genome. In this process, mutations are naturally generated by random integration of mobile DNA elements within a genome. The BonnMu resource is a transposontagged mutant collection, unique in Europe, developed for functional genomics studies in maize (Zea mays L.). In the present project, the mutant repository was expanded by crossing an active Mutator (Mu) line with dent (B73, Co125) and flint (DK105, EP1 and F7) germplasms. By sequencing a total of 8,064 mutagenized BonnMu F2-families we identified 425,924 heritable Mu-insertions affecting 36,612 (83%) of the 44,303 highconfidence gene models of maize. Downstream examination of the heritable Mu-insertions was performed via the automated Mutant-seq Workflow Utility (MuWU). Most of the insertions (94%) were located in coding regions of the genome, mainly in the gene-rich chromosome arms, with 42% in the 5'-untranslated region (UTR). All Mu-insertions and photographs depicting seedling phenotypes from segregating BonnMu F2-families are accessible through the Maize Genetics and Genomics Database (MaizeGDB). In summary, the publicly accessible European BonnMu resource archives insertions for dent and flint lines and is already used worldwide for forward and reverse genetic studies. The genetic diversity represented by the different genetic backgrounds in the BonnMu resource enables the identification of genotype-specific mutations. In the second project, the magenta root dwarf 1 (mrd1) mutant, identified in a forward genetic screening of BonnMu F2-families, was functionally characterized. In comparison to the wild type, the mrd1 mutant accumulates anthocyanins in various root types and exhibit a limited shoot growth under light and dark conditions. The identification of the causal candidate gene was facilitated by the combination of bulked segregant RNA-seq (BSR-seq) and the availability of the sequenced Mu-tagged BonnMu F2-families, which provided a limited number of candidate mutations for the BSR-seq mapping interval. This combinatory approach identified a constitutive photomorphogenesis 9 (cop9) signalosome complex subunit 4 as the causal candidate gene that underlie the mrd1 phenotype. The COP9 signalosome (CSN) is a multiprotein complex initially identified in Arabidopsis thaliana as a repressor of photomorphogenesis. Maize mrd1 mutants show similarities to csn mutants in Arabidopsis such as the constitutive photomorphogenesis, but different patterns of anthocyanin accumulation. PCR- analysis identified mrd1 mutants as homozygous for a Mu-insertion in the mrd1 gene. In a future project, the mrd1 gene could be validated by a gene silencing strategy using RNAi or by a new allele from publicly available mutant resources, such as the continuously expanding BonnMu collection.

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