Abstract
As plants are sessile they need a very efficient system for repairing damage done by external or internal mutagens to their DNA. Mismatch repair (MMR) is one of the systems that maintain genome integrity and prevent homeologous recombination. In all eukaryotes mismatches are recognized by evolutionary conserved MSH proteins often acting as heterodimers, the constant component of which is MSH2. Changes affecting the function of MSH2 gene may induce a ‘mutator’ phenotype and microsatellite instability (MSI), as is demonstrated in MSH2 knock-out and silenced lines of Arabidopsis thaliana. The goal of this study was to screen for ‘mutator’ phenotypes in somatic hybrids between potato cvs. ‘Delikat’ and ‘Désirée’ and MMR deficient Solanum chacoense transformed using antisense (AS) or dominant negative mutant (DN) AtMSH2 genes. The results demonstrate that first generation fusion hybrids have a range of morphological abnormalities caused by uniparental MMR deficiency; these mutant phenotypes include: dwarf or gigantic plants; bushiness; curled, small, large or abnormal leaves; a deterioration in chloroplast structure; small deep-purple tubers and early dehiscent flowers. Forty percent of the viable somatic hybrids planted in a greenhouse, (10 out of 25 genotypes) had mutant phenotypes accompanied by MSI. The majority of the hybrids with ‘mutator’ phenotypes cultured on media containing kanamycin developed roots so sustaining the presence of selectable marker gene nptII, from the initial constructs. Here for the first time, MMR deficiency combined with somatic hybridization, are used to induce new phenotypes in plants, which supports the role of MMR deficiency in increasing introgressions between two related species.
Highlights
Mismatch repair (MMR) is a highly conserved mechanism responsible for maintaining genome stability
The hypothesis tested in this study is that one MMR-deficient parent will increase the number of mutant phenotypes and incidence of homeologous recombination in the resultant somatic hybrids. This is why three transgenic MMR-deficient clones were selected for mesophyll protoplast isolation and fusion with the potato cultivars ‘Delikat’ and ‘Désirée.’ The goals of this study are to: (1) identify abnormal phenotypes in somatic hybrids (SHs) involving MMR-deficient or wild type S. chacoense, in vitro and ex vitro; (2) differentiate between ‘mutator’ phenotypes induced by MMR-deficiency and other abnormal phenotypes caused by somatic fusion or in vitro culture; (3) discuss the role of increased homeologous recombination in inducing the introgression of resistance traits into the potato gene pool; (4) establish the importance of phenotypic variability co-generated by MMR deficiency and somatic hybridization in a wider context by using different tools for breeding potato with many resistant traits
In order to evaluate the effects of MMR deficiency on somatic hybridization it was first important to select the transgenic accession of chc high in leptine carrying the two constructs (Figure 1A) and to confirm the transgenic status of the clones used for protoplast isolation
Summary
Mismatch repair (MMR) is a highly conserved mechanism responsible for maintaining genome stability. The MMR system in plants and other eukaryotes involves many proteins that recognize, excise, and repair the DNA mismatches that occur during DNA replication or due to damage (Bray and West, 2005). MMR Deficient Potato Fusion Hybrids is MSH2, which forms heterodimers with other MSH proteins. These heterodimers recognize mismatches and interact to initiate repair. Recent data indicate that in eukaryotes only 10–15% of MMR events are directly associated with replication and MutSα is able to scan genomes independently. How it does this is unclear (Hombauer et al, 2011). More recent data indicates that in hybrid rice introgressions from the wild species Zizania latifolia induce microsatellite instability, alter MMR activity and result in novel phenotypes (Dong et al, 2013)
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