Fast and Precise: How to Measure Meiotic Crossovers in Arabidopsis

During meiosis, homologous chromosomes (homologs) pair and undergo genetic recombination via assembly and disassembly of the synaptonemal complex. Meiotic recombination is initiated by excess formation of DNA double-strand breaks (DSBs), among which a subset are repaired by reciprocal genetic exchange, called crossovers (COs). COs generate genetic variations across generations, profoundly affecting genetic diversity and breeding. At least one CO between homologs is essential for the first meiotic chromosome segregation, but generally only one and fewer than three inter-homolog COs occur in plants. CO frequency and distribution are biased along chromosomes, suppressed in centromeres, and controlled by pro-CO, anti-CO, and epigenetic factors. Accurate and high-throughput detection of COs is important for our understanding of CO formation and chromosome behavior. Here, we review advanced approaches that enable precise measurement of the location, frequency, and genomic landscapes of COs in plants, with a focus on Arabidopsis thaliana.     

Molecular evolution of the meiotic recombination pathway in mammals

Meiotic recombination shapes evolution and helps to ensure proper chromosome segregation in most species that reproduce sexually. Recombination itself evolves, with species showing considerable divergence in the rate of crossing‐over. However, the genetic basis of this divergence is poorly understood. Recombination events are produced via a complicated, but increasingly well‐described, cellular pathway. We apply a phylogenetic comparative approach to a carefully selected panel of genes involved in the processes leading to crossovers—spanning double‐strand break formation, strand invasion, the crossover/non‐crossover decision, and resolution—to reconstruct the evolution of the recombination pathway in eutherian mammals and identify components of the pathway likely to contribute to divergence between species. Eleven recombination genes, predominantly involved in the stabilization of homologous pairing and the crossover/non‐crossover decision, show evidence of rapid evolution and positive selection across mammals. We highlight TEX11 and associated genes involved in the synaptonemal complex and the early stages of the crossover/non‐crossover decision as candidates for the evolution of recombination rate. Evolutionary comparisons to MLH1 count, a surrogate for the number of crossovers, reveal a positive correlation between genome‐wide recombination rate and the rate of evolution at TEX11 across the mammalian phylogeny. Our results illustrate the power of viewing the evolution of recombination from a pathway perspective.     

Tandem Paired Nicking Promotes Precise Genome Editing with Scarce Interference by p53

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DeepTetrad: high‐throughput image analysis of meiotic tetrads by deep learning in Arabidopsis thaliana

Meiotic crossovers facilitate chromosome segregation and create new combinations of alleles in gametes. Crossover frequency varies along chromosomes and crossover interference limits the coincidence of closely spaced crossovers. Crossovers can be measured by observing the inheritance of linked transgenes expressing different colors of fluorescent protein in Arabidopsis pollen tetrads. Here we establish DeepTetrad, a deep learning‐based image recognition package for pollen tetrad analysis that enables high‐throughput measurements of crossover frequency and interference in individual plants. DeepTetrad will accelerate the genetic dissection of mechanisms that control meiotic recombination.     

Meiotic recombination profiling of interspecific hybrid F1 tomato pollen by linked read sequencing

Genome wide screening of pooled pollen samples from a single interspecific F1 hybrid obtained from a cross between tomato, Solanum lycopersicum and its wild relative, Solanum pimpinellifolium using linked read sequencing of the haploid nuclei, allowed profiling of the crossover (CO) and gene conversion (GC) landscape. We observed a striking overlap between cold regions of CO in the male gametes and our previously established F6 recombinant inbred lines (RILs) population. COs were overrepresented in non‐coding regions in the gene promoter and 5′UTR regions of genes. Poly‐A/T and AT rich motifs were found enriched in 1 kb promoter regions flanking the CO sites. Non‐crossover associated allelic and ectopic GCs were detected in most chromosomes, confirming that besides CO, GC represents also a source for genetic diversity and genome plasticity in tomato. Furthermore, we identified processed break junctions pointing at the involvement of both homology directed and non‐homology directed repair pathways, suggesting a recombination machinery in tomato that is more complex than currently anticipated.     

Four New Members of Foliicolous Green Algae Within the Watanabea Clade (Trebouxiophyceae,Chlorophyta) from China

Members of the Watanabea clade of Trebouxiophyceae are genetically diverse and widely distributed in all kinds of habitats, especially in most terrestrial habitats. Ten new strains of terrestrial algae isolated from the tropical rainforest in China, and four published strains were investigated in this study. Morphological observation and molecular phylogenetic analyses based on the 18S, ITS, rbcL, and tufA genes were used to identify the new strains. Four previously described species were reinvestigated to supplement molecular data and autospores’ morphological photographs. The phylogenetic analyses based on 18S only, the concatenated dataset of 18S and ITS, as well as the concatenated dataset of rbcL and tufA, showed the same phylogenetic positions and relationships of these new strains. According to the phylogenetic analysis and morphological comparisons results, we described these 10 strains as four new members within the Watanabea clade, Polulichloris yunnanensis sp. nov., Polulichloris ovale sp. nov., Massjukichlorella orientale sp. nov., and Massjukichlorella minus sp. nov., and two known species, Massjukichlorella epiphytica, and Mysteriochloris nanningensis. Additionally, we provide strong evidence proving that Phyllosiphon, Mysteriochloris, Polulichloris, and Desertella all reproduce through unequal sized autospores.     

Ligand dynamics controlled reverse spin cross over in bis pyrazolyl pyridine based Fe(II) complex cation with metallodithiolato anions with an example of a ferromagnetic 2:1 cocrystal of mixed Ni(III)/Ni(II) oxidation states

We report here the crystal and molecular structures of three compounds [FeL₂] [Ni(mnt)₂] (1), [FeL₂]₂ [Ni(mnt)₂]₃·2H₂O (2) and [FeL₂] [Cu(mnt)₂]·2CH₃CN (3) where L = 2,6-bis(3,5-dimethylpyrazol-1-ylmethyl)pyridine and mnt = maleonitriledithiolate, and their detailed spectroscopic and magnetic properties using variable temperature Mössbauer, EPR, susceptibility studies, along with room temperature electron spectroscopy for chemical analysis (ESCA) studies. The observed temperature dependant high spin/low spin (HS/LS) ratios of [FeL₂]²⁺ cations in these lattices, exhibiting ‘reverse spin cross-over’ measured unequivocally by Mössbauer, have been interpreted as resulting from differing amount of ‘void space’ in the lattice, a measure of the ease of lattice dynamics originating from ligand L. Differential scanning calorimetric data points this HS/LS transition to order-disorder type of second order phase transitions. While trying to test this lattice dynamics controlled property of [FeL₂]²⁺ cations an unusual behavior of cocrystallization of two planar complex anions of the same type in two different oxidation states, viz. [Ni(mnt)₂]²⁻ and [Ni(mnt)₂]⁻, was observed in [FeL₂]₂ [Ni(mnt₂)]₃, supported by crystallography, ESCA chemical shifts of Ni 2p_3/2 and EPR. The susceptibility data in combination with ESCA chemical shifts of S 2p_3/2 and Ni 2p_3/2 on all the compounds reveal the importance of charge transfer between the two counter ions.     

Recombinase-mediated cassette exchange (RMCE): traditional concepts and current challenges

Traditional DNA transduction routes used for the modification of cellular genomes are subject to unpredictable alterations, as the cell-intrinsic repair machinery may affect both the integrity of the transgene and the recipient locus. These problems are overcome by recombinase-mediated cassette exchange (RMCE) approaches enabling predictable expression patterns by the nondisruptive insertion of a gene cassette at a pre-characterized genomic locus. The destination is marked by a “tag” consisting of two heterospecific recombination target sites (RTs) at the flanks of a selection marker. Provided on a circular donor vector, an analogous cassette encoding the gene of interest can cleanly replace the resident cassette under the influence of a site-specific recombinase. RMCE was first based on the yeast integrase Flp but had to give way to the originally more active phage-derived Cre enzyme. To be effective, both Tyr-recombinases have to be applied at a considerable concentration, which, in the case of Cre, triggers endonucleolytic activities and therefore cellular toxicity. This review addresses the particularities of both recombination routes depending on the structure of the synaptic complex and on improved integrase and RT variants. While the performance of Flp-RMCE can now firmly rely on optimized Flp variants and multiple sets of functional target sites (FRTs), the Cre system suffers from the promiscuity of its RT mutants, which is explained in molecular terms. At present, RMCE enters applications in the stem cell field. Remarkable efforts are noted in the framework of various mouse mutagenesis programs, which, in their first phase, have targeted virtually all genes and now start to shift their emphasis from gene trapping to gene modification.