Molecular Cloning And Function Analysis Of 1R-Myb1 Gene With Single SANT/MYB Domain From Chimonanthus Praecox(L.) Link

Chimonanthus praecox(L.) Link, its English name is wintersweet, which belongs to Calycanthaceae, Chimonanthus Lindl.It blossoms out in the winter, so called wintersweet. Transcription factors are a group of proteins that control cellular processes by regulating the expression of downstream target genes, and they play multi-faceted and pivotal roles in controlling the biological processes of all living organisms.Transcription factors (TFs) implement their functions by binding directly to the promoters of target genes or other proteins which belong to all kinds of transcription factors in a sequence-specific manner to either activate or repress the transcription of downstream target genes.Wintersweet (Chimonanthus praecox Link) is an important species originating from China, where it is cultivated with a long history owing to its important characteristics. They bloom in all kinds of different adversities and their genes related to adversities are rich, as we know many transcription factors take part in the pathway of responsing to environmental stress factors.Comparing oversea research to domestic about wintersweet, we have dominance and get some progress, especially related to biological character, community framework, taxology, investigateing distributing, physiology and biochemistry of flowers and leaves.However, research of Chimonanthus praecox (L.) Link drops behind other plants. In particular, involved molecular biology, reports of Myb gene that contains only one sant domains are few and so are researches as breeding resource.On the basis of Chimonanthus praecox flower cDNA library construction and its ESTs analysis, a 1R-MYB1 protein gene, designated as 1R-Myb1 (GenBank accession number: EU116031), was obtained by sequencing the randomly selected clones. It was showed that the cDNA of 1R-Myb1 was1327bp in length, having no intron and encoding a predicted protein of 318aa.By the way of boifomation, We ananlyse the structure domain model and alignment of lR-Mybl in order to predict its founction. Phosphorylation sites in Ser residue, three sites in Thr residue and four sites in Tyr residue, respectively. Sequence comparisons showed that 1R-Myb1 was a novel transcription factor belonging to MYB family from winterswee, which related to some abiotic stresses.Selecting some organs randomly such as stem, leaf, stamen and gynoecium basing on RT-PCR results, 1R-Myb1 has expressed in these four organs. At the same time, the expression pattern of 1R-Myb1 was analyzed by RT-PCR under some abiotic stresses, such as high temperature, low temperature, high salinity, drought and abscisic acid. It was demonstrated that 1R-Myb1 gene was expressed decreasingly, proposing 1R-Myb1 was negative correlated with abiotic stresses. We speculated 1R-Myb1 was "close-gene". There are three pathways of responsing to environmental stress factors. One is jasmonates signal transduction pathway. The second one is salicy acid (SA). The last one is abscisic acid (ABA). 1R-Myb1 may join in some one of the three pathways and maybe there is crosstalk among the three pathways.By analyzing, 1R-Myb1 participated in ABA pathway mainly, at the sanme time other pathways were secondary. Expression vector was constructed and tranfered into tobacco successfully, for next research by two-hybrid yeast to get down-stream genes of 1R-Myb1.