Cloning of CmAP1 gene from Chrysanthemum morifolium and establishment of its plant expression vector

ObjectiveCmAPETALA1 gene(CmAP1) in Chrysanthemum morifolium was cloned and its plant expres-sion vector was established, in order to provide references for further study on genetic improvement of CmAP1 gene func-tion. MethodCmAP1 gene was cloned from leaf of C. morifolium by method of homology cloning and RT-PCR. The nu-cleotide and amino acid sequence were analyzed by online bioinformatics tool. Expression difference of this gene was deter-mined by real-time fluorescence quantitative PCR. CmAP1 gene was linked to pCAMBIA1300 vector by double enzyme di-gestion to establish plant expression vector. ResultCmAP1 gene(GenBank accession number:KU872999) was obtained by cloning. The open reading frame(ORF) of CmAP1 was 741 bp encoding 246 amino acids. CmAP1 protein was a hy-drophilic protein,and the molecular weight of this protein was about 28.3 kD. Its theoretical isoelectric point (pI) was 8.76. It was predicted that the protein contained ten phosphorylation sites and two potential N-glycosayation sites. Sequence and phylogenic analysis showed that CmAP1 protein was close to C. morifolium CDM111 protein with 98% similarity, which both belonged to euAP1 clade of A type gene in MADS-box gene family. Real fluorescence quantitative PCR analysis indicated that expression of CmAP1 gene was extremely high in buds. The expression level was low in tubiform and lingu-late florets and was in trace expression in vegetative organs. CmAP1 was linked to pCAMBIA1300 vector and the plant expression vector pCAMBIA1300-CmAP1 was established successfully. ConclusionCmAP1 gene plays an important role in growth and development of C. morifolium. The established plant expression vector pCAMBIA1300-CmAP1 is use-ful for further research on changing morphological structure of C. Morifolium by genetic engineering, regulation of flowering and genetic modification.