New Phenotypes of Potato Co-induced by Mismatch Repair Deficiency and Somatic Hybridization

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.

Drought and Saline Stress Tolerance Induced in Somatic Hybrids of Solanum chacoense and Potato Cultivars by Using Mismatch Repair Deficiency

“…MMR-deficient SHs developed significantly more richly (t-test, p < 0.05) and on average 2.14 times longer (t-test, p < 0.05) root systems during severe drought stress than MMR-proficient SHs and BC clones. There was no correlation between drought tolerance and ploidy level, as established in previous analyses [28] and presented for each genotype used in the present experiments (see Figure 2 and Supplementary Tables S1 and S2). The ploidy level determined by flow cytometry and chromosome counting varied in the somatic hybrids with the wild type SHs being predominantly 4× to 5× or 5×, while the MMR-deficient SHs were mostly 4× [28].…”

“…For this study, we used two types of somatic hybrids (SHs) regenerated after mesophyll protoplast electrofusion and hybrid plant regeneration and selection [28]. The first type was obtained by protoplast electrofusion between S. tuberosum cv.…”

“…Delikat (Dk) or Desiree (De) and the high leptine producer (HL), transgenic mismatch repair (MMR) deficient S. chacoense (Table 2). The hybrid status of SHs was confirmed by phenotypic characterization, simple sequence repeat (SSR) marker analysis [28], and by determination of genomic composition using the genomic in situ hybridization (GISH) method (Molnar et al, article in preparation). chacoense used in somatic hybridization, while the number stands for each transgenic clone (e.g., 5 or 11), followed by the number of SH clone; in bold = genotypes used in ex vitro experiment; * = multiple resistance to the biotic (CPB) and abiotic traits (drought, salt).…”

“…chacoense used in somatic hybridization, while the number stands for each transgenic clone (e.g., 5 or 11), followed by the number of SH clone; in bold = genotypes used in ex vitro experiment; * = multiple resistance to the biotic (CPB) and abiotic traits (drought, salt). More details on the SHs characterization can be also found in previous publications [28,32]).…”

“…A function known as anti-recombination prevents the transfer of resistance genes from wild related species into economically important crops. To induce MMR deficiency, S. chacoense was transformed using the AtMSH2 gene in antisense orientation (S) or with a dominant negative mutant sequence (DN) [28].…”