Recombination between homeologous chromosomes, also known as homeologous exchange (HE), plays a significant role in shaping genome structure and gene expression in interspecific hybrids and allopolyploids of several plant species. However, the molecular mechanisms that govern HEs are not well understood. Here, we studied HE events in the progeny of a nascent allotetraploid (genome AADD) derived from two diploid progenitors of hexaploid bread wheat using cytological and whole-genome sequence analyses. In total, 37 HEs were identified and HE junctions were mapped precisely. HEs exhibit typical patterns of homologous recombination hotspots, being biased toward low-copy, subtelomeric regions of chromosome arms and showing association with known recombination hotspot motifs. But, strikingly, while homologous recombination preferentially takes place upstream and downstream of coding regions, HEs are highly enriched within gene bodies, giving rise to novel recombinant transcripts, which in turn are predicted to generate new protein fusion variants. To test whether this is a widespread phenomenon, a dataset of high-resolution HE junctions was analyzed for allopolyploid Brassica, rice, Arabidopsis suecica, banana, and peanut. Intragenic recombination and formation of chimeric genes was detected in HEs of all species and was prominent in most of them. HE thus provides a mechanism for evolutionary novelty in transcript and protein sequences in nascent allopolyploids.

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Breeding of stress-tolerant plants is able to improve crop yield under stress conditions, whereas CRISPR/Cas9 genome editing has been shown to be an efficient way for molecular breeding to improve agronomic traits including stress tolerance in crops. However, genes can be targeted for genome editing to enhance crop abiotic stress tolerance remained largely unidentified. We have previously identified abscisic acid (ABA)-induced transcription repressors (AITRs) as a novel family of transcription factors that are involved in the regulation of ABA signaling, and we found that knockout of the entire family of AITR genes in Arabidopsis enhanced drought and salinity tolerance without fitness costs. Considering that AITRs are conserved in angiosperms, AITRs in crops may be targeted for genome editing to improve abiotic stress tolerance. We report here that mutation of GmAITR genes by CRISPR/Cas9 genome editing leads to enhanced salinity tolerance in soybean. By using quantitative RT-PCR analysis, we found that the expression levels of GmAITRs were increased in response to ABA and salt treatments. Transfection assays in soybean protoplasts show that GmAITRs are nucleus proteins, and have transcriptional repression activities. By using CRISPR/Cas9 to target the six GmAITRs simultaneously, we successfully generated Cas9-free gmaitr36 double and gmaitr23456 quintuple mutants. We found that ABA sensitivity in these mutants was increased. Consistent with this, ABA responses of some ABA signaling key regulator genes in the gmaitr mutants were altered. In both seed germination and seedling growth assays, the gmaitr mutants showed enhanced salt tolerance. Most importantly, enhanced salinity tolerance in the mutant plants was also observed in the field experiments. These results suggest that mutation of GmAITR genes by CRISPR/Cas9 is an efficient way to improve salinity tolerance in soybean.