A designer synthetic chromosome fragment functions in moss

Here, we report the in vivo assembly of chromosomal fragments in the moss Physcomitrium patens, producing phenotypically virtually wild-type lines in which one-third of a chromosomal arm’s coding region is replaced by redesigned, chemically synthesized fragments. By eliminating 58.2% of the sequence, we significantly simplified the genome without discernible phenotypic effects, implying that most transposable elements minimally impact fitness. We also introduced other sequence modifications, such as PCRmark incorporation, gene locus swapping, and stop codon substitution. Despite these substantial changes, the complex epigenetic landscape was correctly reestablished, albeit with some three-dimensional conformation alterations. The synthesis of a partial multicellular eukaryotic chromosome arm lays the foundation for the synthetic moss genome project (SynMoss) and paves the way for genome synthesis in multicellular organisms.