The characterisation of transcriptome from different development stages of Rafflesia cantleyi floral buds

Abstract

Rafflesia cantleyi (R. cantleyi) is a specialised holoparasitic plant with dramatic morphological modifications. It has a reduced vegetative structure, which only appears as a flower and it possesses an unusual life cycle, where its floral bud development takes up to nine months. However, little information is available on the underlying mechanisms that govern flower development in this organism. To gain a global perspective on the flower development in R. cantleyi, transcriptome data were generated from three developmental stages of its floral buds, representing the early (FBS1), mid (FBS2) and advanced (FBS3) developmental stages that were used to profile the floral bud transcriptomes and identify differentially expressed transcripts (DEGs) between the three floral bud stages (FBS1 vs. FBS2, FBS2 vs. FBS3, and FBS1 vs. FBS3). A total of 89 690 transcripts were obtained by de novo assembly of 91.63 million clean paired-end reads through the Trinity analysis pipeline. A total of 2312 and 3961 DEGs were identified for FBS1-FBS2 and FBS2-FBS3 comparisons, respectively. A total of 972 and 1769 transcripts were up-regulated; while 1340 and 2192 transcripts were down-regulated in FBS1-FBS2 and FBS2-FBS3, respectively. In the FBS1-FBS3 comparisons, 4642 DEGs were identified with 1999 up-regulated and 2 643 down-regulated. Using K-mean clustering, DEGs in 12 co-expression clusters with different patterns were identified. The centroid values of each cluster which show the expression patterns of each cluster were used to compare the relationships between clusters, and three major groups were identified. These three major group showed correlations to three developmental stages. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) enrichment analyses have revealed that DEGs are involved in hormone signal transduction, light signalling, and the specification of floral organ identity. Among these DEGs are transcription factor families involved in flower development, such as ARF, MADS-box, bHLH, and MYB families. Furthermore, DEGs associated with hormone auxin (IAA), cytokinin (CK), gibberellic acid (GA), jasmonic acid (JA) and as well as flowering pathways such as photoperiod and autonomous pathways were identified. The phytohormone-related DEGs suggest similar roles of phytohormones during flowering process and highlight the possible role of GA and JA in R. cantleyi flowering. A total of 628 transcripts exhibiting sequence similarities to Arabidopsis flowering-related genes include SVP, LFY, and members of ABCE model genes were differentially expressed during flower development. Real-time quantitative polymerase chain reaction (RT-qPCR) analysis was performed to validate the expression of 12 candidate genes. In addition, floral organ-specific expression patterns of ABCE model genes showed the relationship between ABCE model genes expression and floral morphology in R. cantleyi. Based on this, one ABCE model was proposed. The results presented in this study represent a comprehensive genetic reference for future studies of floral bud development and flowering process in R. cantleyi and other related species.