Transcriptome analysis of hormone-induced embryoid formation in un-pollinated pumpkin ovary

ZHANG Zixia; LIU Zefa; TANG Yulong; LIANG Zhirui; ZHU Hailong; CHEN Ziyao

doi:10.16861/j.cnki.zggc.2025.0564

Abstract：Induction of haploid by in vitro culture of unpollinated ovary is one of the important ways of genetics and breeding.In order to elucidate the molecular mechanism underlying hormone induced embryoid formation from unpolli-nated pumpkin ovary,ZJ-13-2(Chinese pumpkin×Indian pumpkin)was used as the material in this study.Auxin and cyto-kinin were used to induce embryoids at the differentiation stage.The results showed that the induction rate of embryoids in the medium supplemented with cytokinin and auxin(1.0 mg·L-16-BA+4.0 mg·L-12,4-D+0.5 mg·L-1 NAA)was sig-nificantly higher than that in the medium supplemented with cytokinin only(1.0 mg·L-16-BA),which were 32.1%and 13.8%,respectively.Transcriptome sequencing of embryoids from both treatment groups was performed to analyze differ-entially expressed genes(DEGs)influencing embryoid formation under hormone synergy.A total of 1197 DEGs were iden-tified,comprising 675 up-regulated and 522 down-regulated genes.Go and KEGG enrichment analyses revealed that these DEGs were mainly enriched in plant hormone signal transduction,protein processing in endoplasmic reticulum,starch and sucrose metabolism,and secondary metabolite synthesis pathways.Further analysis identified 441 transcription factors(TFs)belonging to 47 different TF families.Key TFs,including CRYAA,CRYAB,HSP20,SKP1,BKI1,and BZR2 exhibited significant expression changes,suggesting their potential regulatory roles in the embryogenic process of this pumpkin variety.Additionally,HSF family members were found to interact with other transcription factors such as WRKY,bZIP and MYB,collectively regulating the expression of heat stress responsvie genes.This complex regulatory network not only plays a key role in heat stress response,but also provides actionable molecular markers and genetic re-sources for constructing an efficient haploid induction system in pumpkin,thereby accelerating the fixation of important traits and the overall breeding process.