Allodiploid hybridization, loss-of-heterozygosity and aneuploidy in the green alga Auxenochlorella , an emerging model for discovery research and bioengineering

Auxenochlorella are diploid oleaginous green algae that are highly amenable to genetic transformation by homologous recombination, providing great potential for discovery research and bioengineering. Vegetatively diploid organisms experience specific evolutionary phenomena, including allodiploid hybridization, mitotic recombination, loss-of-heterozygosity and aneuploidy; however, studies of these forces have mostly focused on fungi. Here, we present a telomere-to-telomere phased diploid genome assembly of Auxenochlorella UTEX 250-A, revealing a streamlined nuclear genome architecture with a haploid length of 22 Mb. We demonstrate that UTEX 250-A is an allodiploid hybrid via sequencing of Auxenochlorella protothecoides and Auxenochlorella symbiontica strains, two close relatives differentiated by extensive chromosomal rearrangements. The UTEX 250-A haplotypes are a mosaic of each parental species following mitotic recombination, and two chromosomes are trisomic, which we confirm via allele-specific transformation of a trisomic chromosome. Loss-of-heterozygosity events are likely pervasive across Auxenochlorella due to mitotic recombination and transient aneuploidy, and can evolve rapidly in laboratory culture. We present high-quality structural annotations, yielding ∼7,500 genes per haplotype. Auxenochlorella have experienced extensive gene family loss and reduction, including core photosynthesis genes. Nevertheless, a minimal set of genes required for sexual reproduction is potentially present. We demonstrate periodic adenine and cytosine methylation around promoters and across gene bodies. Finally, ∼10% of genes, especially those involved in DNA repair and sex, overlap antisense long noncoding RNAs, potentially representing a regulatory mechanism. These results demonstrate the generality of several evolutionary forces associated with vegetative diploidy, and provide a genomic foundation for the use of Auxenochlorella as a reference organism.