Break-induced replication underlies formation of inverted triplications and generates unexpected diversity in haplotype structures.

Background: The duplication-triplication/inverted-duplication (DUP-TRP/INV-DUP) structure is a type of complex genomic rearrangement (CGR) hypothesized to result from replicative repair of DNA due to replication fork collapse. It is often mediated by a pair of inverted low-copy repeats (LCR) followed by iterative template switches resulting in at least two breakpoint junctions in cis . Although it has been identified as an important mutation signature of pathogenicity for genomic disorders and cancer genomes, its architecture remains unresolved and is predicted to display at least four structural variation (SV) haplotypes. Results: Here we studied the genomic architecture of DUP-TRP/INV-DUP by investigating the genomic DNA of 24 patients with neurodevelopmental disorders identified by array comparative genomic hybridization (aCGH) on whom we found evidence for the existence of 4 out of 4 predicted SV haplotypes. Using a combination of short-read genome sequencing (GS), long- read GS, optical genome mapping and StrandSeq the haplotype structure was resolved in 18 samples. This approach refined the point of template switching between inverted LCRs in 4 samples revealing a DNA segment of ∼2.2-5.5 kb of 100% nucleotide similarity. A prediction model was developed to infer the LCR used to mediate the non-allelic homology repair. Conclusions: These data provide experimental evidence supporting the hypothesis that inverted LCRs act as a recombinant substrate in replication-based repair mechanisms. Such inverted repeats are particularly relevant for formation of copy-number associated inversions, including the DUP-TRP/INV-DUP structures. Moreover, this type of CGR can result in multiple conformers which contributes to generate diverse SV haplotypes in susceptible loci .

The coexpression of two desaturases provides an optimized reduction of saturates in camelina oil.

Reducing the saturate content of vegetable oils is key to increasing their utility and adoption as a feedstock for the production of biofuels. Expression of either the FAT5 16 : 0-CoA desaturase from Caenorhabditis elegans, or an engineered cyanobacterial 16 : 0/18 : 0-glycerolipid desaturase, DES9*, in seeds of Arabidopsis (Arabidopsis thaliana) substantially lowered oil saturates. However, because pathway fluxes and regulation of oil synthesis are known to differ across species, translating this transgene technology from the model plant to crop species requires additional investigation. In the work reported here, we found that high expression of FAT5 in seeds of camelina (Camelina sativa) provided only a moderate decrease in saturates, from 12.9% of total oil fatty acids in untransformed controls to 8.6%. Expression of DES9* reduced saturates to 4.6%, but compromised seed physiology and oil content. However, the coexpression of the two desaturases together cooperatively reduced saturates to only 4.0%, less than one-third of the level in the parental line, without compromising oil yield or seedling germination and establishment. Our successful lowering of oil saturates in camelina identifies strategies that can now be integrated with genetic engineering approaches that reduce polyunsaturates to provide optimized oil composition for biofuels in camelina and other oil seed crops.