Harnessing clonal gametes in hybrid crops to engineer polyploid genomes.

Heterosis boosts crop yield; however, harnessing additional progressive heterosis in polyploids is challenging for breeders. We bioengineered a 'mitosis instead of meiosis' (MiMe) system that generates unreduced, clonal gametes in three hybrid tomato genotypes and used it to establish polyploid genome design. Through the hybridization of MiMe hybrids, we generated '4-haplotype' plants that encompassed the complete genetics of their four inbred grandparents, providing a blueprint for exploiting polyploidy in crops.

Technology-driven approaches for meiosis research in tomato and wild relatives.

Meiosis is a specialized cell division during reproduction where one round of chromosomal replication is followed by genetic recombination and two rounds of segregation to generate recombined, ploidy-reduced spores. Meiosis is crucial to the generation of new allelic combinations in natural populations and artificial breeding programs. Several plant species are used in meiosis research including the cultivated tomato (Solanum lycopersicum) which is a globally important crop species. Here we outline the unique combination of attributes that make tomato a powerful model system for meiosis research. These include the well-characterized behavior of chromosomes during tomato meiosis, readily available genomics resources, capacity for genome editing, clonal propagation techniques, lack of recent polyploidy and the possibility to generate hybrids with twelve related wild species. We propose that further exploitation of genome bioinformatics, genome editing and artificial intelligence in tomato will help advance the field of plant meiosis research. Ultimately this will help address emerging themes including the evolution of meiosis, how recombination landscapes are determined, and the effect of temperature on meiosis.

A chromosome scale tomato genome built from complementary PacBio and Nanopore sequences alone reveals extensive linkage drag during breeding.

The assembly and scaffolding of plant crop genomes facilitate the characterization of genetically diverse cultivated and wild germplasm. The cultivated tomato (Solanum lycopersicum) has been improved through the introgression of genetic material from related wild species, including resistance to pandemic strains of tobacco mosaic virus (TMV) from Solanum peruvianum. Here we applied PacBio HiFi and ONT Nanopore sequencing to develop independent, highly contiguous and complementary assemblies of an inbred TMV-resistant tomato variety. We show specific examples of how HiFi and ONT datasets can complement one another to improve assembly contiguity. We merged the HiFi and ONT assemblies to generate a long-read-only assembly where all 12 chromosomes were represented as 12 contiguous sequences (N50 = 68.5 Mbp). This chromosome scale assembly did not require scaffolding using an orthogonal data type. The merged assembly was validated by chromosome conformation capture data and is highly consistent with previous tomato genome assemblies that made use of genetic maps and Hi-C for scaffolding. Our long-read-only assembly reveals that a complex series of structural variants linked to the TMV resistance gene likely contributed to linkage drag of a 64.1-Mbp region of the S. peruvianum genome during tomato breeding. Through marker studies and ONT-based comprehensive haplotyping we show that this minimal introgression region is present in six cultivated tomato hybrid varieties developed in three commercial breeding programs. Our results suggest that complementary long read technologies can facilitate the rapid generation of near-complete genome sequences.

Epigenetic activation of meiotic recombination near Arabidopsis thaliana centromeres via loss of H3K9me2 and non-CG DNA methylation.

Eukaryotic centromeres contain the kinetochore, which connects chromosomes to the spindle allowing segregation. During meiosis, centromeres are suppressed for inter-homolog crossover, as recombination in these regions can cause chromosome missegregation and aneuploidy. Plant centromeres are surrounded by transposon-dense pericentromeric heterochromatin that is epigenetically silenced by histone 3 lysine 9 dimethylation (H3K9me2), and DNA methylation in CG and non-CG sequence contexts. However, the role of these chromatin modifications in control of meiotic recombination in the pericentromeres is not fully understood. Here, we show that disruption of Arabidopsis thaliana H3K9me2 and non-CG DNA methylation pathways, for example, via mutation of the H3K9 methyltransferase genes KYP/SUVH4 SUVH5 SUVH6, or the CHG DNA methyltransferase gene CMT3, increases meiotic recombination in proximity to the centromeres. Using immunocytological detection of MLH1 foci and genotyping by sequencing of recombinant plants, we observe that H3K9me2 and non-CG DNA methylation pathway mutants show increased pericentromeric crossovers. Increased pericentromeric recombination in H3K9me2/non-CG mutants occurs in hybrid and inbred backgrounds and likely involves contributions from both the interfering and noninterfering crossover repair pathways. We also show that meiotic DNA double-strand breaks (DSBs) increase in H3K9me2/non-CG mutants within the pericentromeres, via purification and sequencing of SPO11-1-oligonucleotides. Therefore, H3K9me2 and non-CG DNA methylation exert a repressive effect on both meiotic DSB and crossover formation in plant pericentromeric heterochromatin. Our results may account for selection of enhancer trap Dissociation (Ds) transposons into the CMT3 gene by recombination with proximal transposon launch-pads.