Cloning and expression analysis of GRF genes related to spur development in Impatiens uliginosa Franch.

GRF genes related to spur development in Impatiens uliginosa Franch. were cloned using reverse transcription polymerase chain reaction (RT-PCR), with homology and phylogenetic analysis of protein sequences then investigated using DNAMAN and MEGA. In addition, the spatiotemporal expression patterns of GRF genes were investigated by quantitative RT-PCR(qRT-PCR). Results showed that two GRF genes of I. uliginosa were successfully cloned, whose full-length cDNA sequences were 1137 bp and 903 bp, encoding 378 amino acids (aa) and 300 aa and named IuGRF1 and IuGRF2, respectively. Both contained the QLQ and WRC structural domains. The GRF genes of I. uliginosa showed low homology with GRF genes from other species, except for the conserved domains, which may be correlated with the diversity of the C-terminal domain in GRF genes. Based on phylogenetic analysis, IuGRF1 and IuGRF2 were separated into different branches. The qRT-PCR results showed that the expression of IuGRF1 was the highest in the spur cup at both the bud and blooming stages but decreased in the blade with the development of the spur. The expression of IuGRF2 was the highest in the blade at the bud stage but declined gradually with spur growth and was low in the spur cup. The expression levels of IuGRF1 and IuGRF2 were the highest in the cup curve at the beginning of the flowering stage but decreased gradually in the cup mucro and curve with spur development and increased gradually in the cup base. Based on the above results, we speculated that the two GRF genes in I. uliginosa were paralogous, with IuGRF1 playing a major role in spur cup development and IuGRF2 playing a major role in blade development.