Objective The research on the regulation of NF-Y nuclear factor on cone development in conifers has not been reported yet. Through the cloning, expression characteristics and functional analysis of PtNF-YC1 gene of Pinus tabuliformis, it provides a basis for the functional study of conifer NF-Y gene family in the reproductive development of conifers.
Method (1) The relationship between PtNF-YC1 and Arabidopsis thaliana NF-YC subfamily proteins was analyzed by phylogenetic tree. (2) Tobacco was transiently transformed to detect the subcellular localization of PtNF-YC1. (3) The expression characteristics of PtNF-YC1 in different tissues of P. tabuliformis were analyzed based on transcriptome data. (4) PtNF-YC1 was heterologously transformed into Arabidopsis thaliana , and the flowering time of different transgenic Arabidopsis thaliana lines under long-day and short-day was compared. The transcriptome sequencing of each transgenic line under long-day was performed to screen the genes related to PtNF-YC1 regulating flowering. (5) The protein interaction between PtNF-YC1 and candidate proteins was verified by Y2H and BiFC.
Result The open reading frame of PtNF-YC1 was 897 bp, which encoded 299 amino acids, had a typical NF-YC conserved domain, and a close relationship with the homologous genes of AtNF-YC3/4/9. The subcellular localization showed that PtNF-YC1 was localized in the nucleus and cytoplasm. The analysis of expression patterns in different tissues showed that PtNF-YC1 could be expressed in needles, vegetative buds, male and female cones and roots, but the expression abundance was the highest in male cones and stems. PtNF-YC1 heterologous transformation of Arabidopsis thaliana delayed flowering under short day. It was proved that PtNF-YC1 interacted with PtCOL5 through Y2H and BiFC.
Conclusion PtNF-YC1 has the function of regulating flowering time and is a candidate gene for inducing cone development through photoperiodic pathway of P. tabuliformis.