Characterization of Flowering-associated Genes in Citrus and Applications for Reducing Breeding Time

Many commercial citrus varieties have a long juvenile period, often taking over 6 years to reach reproductive maturity. This long generation time hinders citrus breeders from quickly developing new varieties with superior genetics. Therefore, early flowering is a desired characteristic for breeding new cultivars and elucidating the molecular mechanisms to efficiently induce early flowering is a useful biotechnology for citrus breeders and producers. However, the factors controlling flowering in citrus are not fully understood. This study focuses on a protein family with a specific phosphatidylethanolamine binding domain (PEBP domain). This highly conserved gene family is part of an intricate signaling network controlling floral transition and has been well characterized in Arabidopsis thaliana, but has not been fully studied in citrus. In Arabidopsis this gene family has 6 members, FLOWERING LOCUS T (FT), TERMINAL FLOWER1 (TFL1), MOTHER OF FT (MFT), BROTHER OF FT (BFT), ARABIDOPSIS THALIANA CENTRADIALIS HOMOLOGUE (ATC), and TWIN SISTER OF FT (TSF). Although these genes share high homology at the deduced amino acid sequence level, some of these genes repress while others promote flowering. To identify the phylogenetic relatedness between proteins we compared homology of the Arabidopsis proteins with those of Citrus clementina and C. sinensis. This led us to identify 8 PEBP gene family members in C. clementina and 10 in C. sinensis. Phylogenetic trees were constructed to compare relatedness between proteins. Additionally, we identified early and late flowering lines from a segregating F₂ mapping population derived from crosses between ‘Pineapple’ Sweet Orange (C. sinensis) and Eremocitrus glauca. To understand the interplay between the different flowering genes in citrus, we used quantitative RT-PCR to analyze transcript levels of flowering-associated genes and determined which of the genes coding PEBPs are differentially expressed in the leaves of early and late flowering individuals. Data from this study will be utilized to identify candidates for artificial manipulation of gene expression and induce early flowering in citrus with the overall goal of reducing breeding time.