Isolation and expression analysis of a reference gene: ClTUA of Chrysanthemum lavandulifolium.

Selection and validation of reference genes constitute a key point for gene expression analysis of Chrysanthemum lavandulifolium. An EST encoding α-tubulin from C. lavandulifolium was identified in our previous study. By using RACE approach, we isolated the full length sequence of this gene and designated it as ClTUA. ClTUA contained an open reading frame of 1 580 bp that encoded a protein of 432 amino acids. Phylogenetic analysis using MEGA4.0 software indicated that ClTUA belonged to class Ⅰ of α-tubulin genes. To assess the usability of ClTUA as a reference gene, we compared the expression patterns of ClTUA with two other housekeeping genes, ClActin and 26S rRNA under salt, drought, cold, heat, ABA, and SA treatments as well as in different developmental stages. Our results showed that the transcript levels of ClActin and 26S rRNA were stable under abiotic treatments but varied in different developmental stages, while that of ClTUA was expressed consistently in all samples. The data suggest that ClTUA is more suitable than ClActin and 26S rRNA as a reference gene for the analysis of gene expression patterns under abiotic stress and in different growth stages in C. lavandulifolium.