Molecular cloning and functional analysis of Chinese bayberry MrSPL4 that enhances growth and flowering in transgenic tobacco

Chinese bayberry ( Myrica rubra ) is an important tree in South China, with its fruit being of nutritional and high economic value. The early maturing varieties of Chinese bayberry are characterized by early flowering. In this study, early ripening (ZJ), medium ripening (BQ) and late ripening (DK) varieties were used as test materials, while young leaves in the floral bud morphological differentiation period were selected for transcriptome sequencing. A total of 4,538 differentially expressed genes were detected. Based on clustering analysis and comparisons with genes reportedly related to flowering in Arabidopsis thaliana , 25 homologous genes were identified. Of these, one was down-regulated in DK but up-regulated in ZJ and BQ, with the results also reflected in the relative expression determined by qRT-PCR. The gene contained the SBP domain and the target site of miR156, named MrSPL4 . Primers were designed to clone the gene whose total length and the length of the CDS sequence were 1,664 bp and 555 bp respectively. An overexpression vector (35S:: 35S:: MrSPL4 -pCambia2301-KY) was then constructed and successfully transfected into tobacco to obtain MrSPL4 -positive plants. In this case, the relative expression of MrSPL4 was up regulated by 3,862.0-5,938.4 times. The height of positive plants was also significantly higher than that of wild-type (WT) plants, with the bud stage occurring 12 days earlier. Altogether, the results indicated that the gene influenced growth and flowering, which provided a theoretical basis for the regulation of flowering in Chinese bayberry.