关键词: 芥菜, SVP, 酵母双杂交, 开花调控

Abstract:

SHORT VEGETATIVE PHASE（SVP）was a key regulatory factor in flowering-time control of Brassica juncea. In order to clarify the expression characteristics of SVP gene and the mediation sites of protein interactions between SVP with FLC in flowering pathways，we cloned SVP gene in‘Qingyejie’germplasm of Brassica juncea. Expression analysis of qRT-PCR revealed that SVP gene expressed in leaves as well as shoot apexes in the flowering pathways of vernalization and long-day photoperiod.   SVP gene expressed at very low level（0.56 in stem tip and 0.35 in leaf blade） at early stage of vegetative phase. However，it significantly increased at early stage of the reproductive phase（The relative expression values in the vernalization pathway are 0.60 in stem tip and 1.27 in leaf blade，respectively. However，the values in the photoperiod pathway are 0.49 in stem tip and 1.42 in leaf blade）. In stem tips，the expression of SVP was more sensitive to vernalization than that of long-day photoperiod. Contrarily，the sensitivity of SVP gene expression in leaves was exactly reversed. Yeast two-hybrid experiments and β-galactosidase activity assays showed that FLC₂ retained protein interactions withI-domain mutantof SVP^E90L as well as K-domain mutants of SVP^K104C and SVP^H106I. However，the interacting strength of SVP/FLC₂ was weakened by each of the three mutation sites above mentioned. Another K-domain mutant of SVP^R137L failed to interact with FLC₂ but still interacted with FLC₁，FLC₃，FLC₄ and FLC₅. It suggested that the interaction of SVP/FLC₂ was specifically regulated by the 137th amino acid site of SVP. Sequence comparison showed that two different genes of FLC₄ and FLC_{5 encoded the same FLC protein，which had only one variable amino acid site compared with FLC3. However，there are respectively 28，19 and 18 variable amino acid sites in FLC1，FLC3 and FLC4-5 compared with FLC2. Additionally，FLC2 had 11 unique amino acid sites which were different from any of other FLC family members such as FLC1，FLC3，FLC4 or FLC5. Hence，we speculated that the variable sites between FLC2 and other FLC members probably contributed to the specific regulation of SVP^R137L/FLC2.}in B. juncea

Key words:  Brassica juncea, SVP, yeast two-hybrid system, flowering regulation