| Grapevine(Vitis)is one most popular fruit tree in the world and it plays an important role in the agricultural economy of China.The floral formation of grapevine is characterized as easy,and it can also easily happen at top nodes of grapevine canes.For many grape varieties,the winter buds growing at nodes from the basal to the top one of the cane can differentiate into floral buds,and blossom and yield the next year.Although their floral differentiating periods are quite different,their flowering and fruiting time are quite similar,which indicates the flora-differentiating rates of buds on all the node of same canes are different at first,however,reach to synchronization eventually.The mechanism underlying this phenomenon has not been focused and studied.Based on the mentioned above,in this study,a easily flora-differentiating grapevine cultivar 'Fujiminori' was used as plant material to characterize the phenomenon in grape and the expression patterns of the flowering-related genes in the buds at different cane nodes,from which the characteristic of early or late flora-differentiation,differentiating rates of the buds at different cane nodes,and the last synchronic-development phenomenon were briefly uncovered.Moreover,the pathways of the flowering were also verified.The main results went as follows:To investigate the proceeding of development of flower buds,the phonological of buds development at different nodes were observed in 'Fujiminori' by using cutting back and debudding at different development period.The results showed that buds at different nodes of the treated and control branches can form flowers,he developments of the buds at different nodes of grapevine branch are different.The first bud began to form and develop from late April or early May,and the formation time of the last bud at the top was in late September.The qualities of the central buds on a branch are higher than those buds at upper and lower nodes of the same branches.The development time of the lower nodes are five months longer than the upper nodes,but the flowering time of them are almost at the same in the second season.This suggests that there is gradually synchronization develop process in buds in different nodes.Based on the nucleotide sequences deposited in Genbank,seven pairs of primers were designed according to the sequence of important genes(VvAG,VvAP1,VvAP3,VvFT,VvFLC,VvSOCl,VvFUL)involved in grapevine floral development.Their homologues sequences were amplified in grapevine cultivar 'Fujiminori',and the fragments flanking open reading frame were coloned and sequenced,respectively.The deduced amino acid sequences of the 7 genes were aligned and blasted with their corresponding sequence,and seven phylogenetic trees were constructed.The results showed that the 7 genes involved in floral development in 'Fujiminori' and woody plants have high similarity with the genes in Arabidopsis.This result indicated that the 7 genes were evolved randomly in 'Fujiminori'.Using gene sequences involved in floral development pathway of Arabidopsis as the probes,homologous EST sequences were searched against the grape EST database.By using cDNAs from different grape organs at various developing stages,half-qRT-PCR were preformed to analyze the expression of these genes,and floral development pathway were forecasted in grapevine.The study verified that most of the genes participating in Arabidopsis flowering pathways were also present in the grape and the expression of these genes exists temporal and spatial difference.To reveal the mechanism of the period and speed of buds development on different nodes of grape branches,expression pattern of 8 floral genes involved in the floral development were alalyzed in the buds at different nodes of 'Fujiminori' in the whole year and analyzed by real-time quantitative PCR.The result showed that the buds remain strong physiological activities in the dormancy period.The expression levels of these floral genes in the buds at different nodes are different.The expression of VvFT is low at the dormancy period and reaches to the peak at budding period.VvSOC1 has two expression peak,and time period between two peaks becomes shorter with the buds rise.Floral genes VvAP1? VvAP2?VvAP3?VvFUL and VvAG showed different expression pattern.VvAP1 and VvAP3 have two expression peak,while VvAP2?VvFUL and VvAG have one expression peak.VvFLC keeps high expression level during the bud developing period,and the expression peak appears in September to October.After the buds mature,the expression starts to decrease.The lengths of high level expression period of floral genes in the buds are different from nodes to nodes.The development time of upper buds is short before bud dormancy,but their active differentiation period is relatively long.The lower buds also had higher or similar levels of gene expression than the central buds in the winter dormant period.Genes from different nodes appear the expression peak at different time.Most of the genes appear the expression peak early will turn to low expression levels more quickly,suggesting that although the peak period of floral development of lower and upper buds are relatively late,development will continue after the peak period,and ultimately lead to the floral differentiation simultaneously in lower,middle and upper nodes.The results predict that the dormancy is the critical period of synchronization of flower buds growing at different nodes of grapevine cane.In order to reveal the biological networks underlying the synchronization character of flower buds differentiation,RNA sequence was conducted by using buds samples from different nodes of 'Fujiminori' grapevine cane at initial dormancy period.A total of 28161327,28808716 and 27941132 clean reads were obtained from bottom,middle and top nodes by RNA sequence,respectively.By pairwise comparisons,more differentially expressed genes(2639)were observed between top and bottom buds than those(1564)between top and middle buds,than those(398)between middle and bottom buds.Among the differentially expressed genes,more up-regulated genes were observed in top buds than middle,than bottom.By GO analysis,differentially expressed genes were classified into 23 of Biology Process,11 of Cell Component and 11 of Molecular Function,mainly in Cellular Process,Metabolic Process,Response to Stimulus,Biological Regulation,Localization,Establishment of Localization,Cell,Cell Part,Organelle,Organelle Part,Macromolecular Complex,Binding,Catalytic Activity,and Structural Molecule Activity.One hundred and one genes concerned in floral development pathway were found in transcriptome data.Eighteen out of 101 were differentially expressed in bottom,middle and top buds by pair wised comparison,and 14 out of the 18 genes were up-regulated at upper nodes,which could explain synchronization of flower buds growing at different nodes.The 18 differentially expressed genes were concerned in vernalization pathway,photoperiod pathway and gibberellin pathway.The transcriptome data accumulated in this study could provide valuable reference for further investigation of floral development in 'Fujiminori'. |
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