Gene Expression Dynamics During Somatic Embryogenesis in Woody Plant Species

Abstract

Somatic embryogenesis is the only cloning route capable of delivering conifer propagation at a scale useful to
forestry, yet the low and unpredictable conversion of proembryogenic masses into mature, germinable embryos
remains the single largest obstacle to routine deployment. We used RNA sequencing to track gene expression
across five developmentally defined stages of somatic embryogenesis in Norway spruce (Picea abies), from
proliferating proembryogenic masses through early and late somatic embryos to the desiccation-tolerant
maturation stage, comparing two embryogenic cell lines with contrasting maturation efficiency (line H selected for
high conversion, line L for persistently low conversion despite repeated maturation attempts). Differential
expression analysis identified 6,847 genes changing significantly across the developmental series in line H,
organised into six temporal clusters by weighted gene co-expression network analysis. The transition from
proembryogenic mass proliferation to early embryo patterning coincided with sharp induction of LEAFY
COTYLEDON, BABY BOOM and WOX2 orthologues alongside a marked shift in auxin-responsive gene
expression consistent with establishment of the apical-basal polarity axis. Line L showed a substantially blunted
and delayed version of this transcriptional switch, with LEAFY COTYLEDON induction reduced by roughly 70% at
the equivalent developmental timepoint and persistent expression of proliferation-associated cyclin genes into
stages where line H had already committed to embryo patterning, suggesting a failure to exit the proembryogenic
proliferative state cleanly rather than a defect specific to later maturation physiology. Late-stage maturation was
dominated in both lines by late embryogenesis abundant protein and storage protein transcripts, but line L
reached only 40-55% of line H's peak expression for the core LEA gene set. These results place the
proembryogenic-to-embryo transition, rather than desiccation tolerance acquisition itself, as the developmental
bottleneck most clearly distinguishing high- and low-conversion embryogenic lines, and they identify a compact
panel of marker transcripts that could be used to triage new embryogenic lines before committing them to lengthy
maturation protocols.