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以‘秋发1号’和‘洛阳红’牡丹(Paeonia suffruticosa)为试验材料,采用RT-PCR的方法从花芽中克隆得到1个隐花色素基因Cryptochrome 2,其ORF全长为1902 bp,编码633个氨基酸,基因登录号为KP982893。序列比对和结构域分析表明,此蛋白包含1个PHR和1个CCT结构域,与拟南芥中AtCRY2最为相似,将其命名为PsCRY2。系统进化树分析表明,PsCRY2与甜橙(Citrus sinensis)CsCRY2亲缘关系最近。实时荧光定量PCR表明,PsCRY2在牡丹的不同组织器官中均有表达。其中,成年植株的花芽以及种子胚的表达量最高,幼苗根、茎、叶次之。在整个花芽分化不同时期,PsCRY2的表达呈现较高的表达水平;在花蕾发育不同时期,PsCRY2的表达呈现先降低再升高而后有降低的趋势,大风铃期表达量最高,推测PsCRY2在花芽分化和花蕾发育的过程中均起到重要作用。在‘洛阳红’和‘秋发1号’牡丹春季花芽发育过程中,PsCRY2的表达均呈升高的趋势,‘秋发1号’显著高于‘洛阳红’。不同光周期条件下,PsCRY2表达稍有不同。与长日照条件相比,短日照条件下PsCRY2在现蕾期和小风铃期的表达量均下降。
Cryptochrome 2 (Cryptochrome 2) was cloned from flower buds by RT-PCR using ’Paeonia suffrutosa’ and ’Qiufa 1’ and ’Luoyan 1’. The full length ORF of Cryptochrome 2 was 1902 bp, 633 amino acids, gene accession number KP982893. Sequence alignment and domain analysis showed that this protein contains 1 PHR and 1 CCT domain, which is most similar to AtCRY2 in Arabidopsis and named as PsCRY2. Phylogenetic tree analysis showed that PsCRY2 has the closest relationship with CsCRY2 of Citrus sinensis. Real-time PCR showed that PsCRY2 was expressed in different tissues and organs of Peony. Among them, the flower buds and seed embryos of adult plants had the highest expression, followed by the roots, stems and leaves of the seedlings. At different stages of flower bud differentiation, the expression of PsCRY2 showed a higher expression level. At different stages of bud development, the expression of PsCRY2 decreased first and then increased and then decreased, and the expression of PsCRY2 was the highest during the flowering stage. And bud development play an important role in the process. During the flower bud development of ’Luoyanghong’ and ’Qiufa 1’ in spring, the expression of PsCRY2 showed an increasing trend, and ’Qiufa 1’ was significantly higher than that of ’Luoyanghong’. The expression of PsCRY2 was slightly different under different photoperiod. Compared with the long-day condition, the expression of PsCRY2 in short-day condition decreased in budding stage and in small wind-bell stage.