Genome-wide Analysis Of Zizania Latifolia D-type Cyclin And Effect Of CKX Gene Expression In T-type Strains Of U.esculenta P.Henn

Plant growth and development through somatic cell division into special functional organs.Cyclin plays an important role in promoting cell cycle progression.Among them,CycD-related kinases mainly regulate phosphorylation of retinoblastoma(RBR)proteins to regulate G1/S conversion.And plant hormones such as auxins and cytokinins also play an important role in the regulation of cell cycle progression.There have been many studies on cyclins so far,but cyclins have not been studied for monocotyledonous Jiaobai plants.The phylogenetic tree and proteomics analysis of Jiaobai CycD was performed by bioinformatics software.According to the published transcriptome data of HSD and Zhejiao 2,and this laboratory has also selected Jiaobai of different periods Jiaobai to perform transcriptome sequencing.According to the results of the sequencing,We analyze the amount of Jiaobai expression.Finally we selected key genes using qRT-PCR for gene expression verification and in situ hybridization experiment.Since cytokinin is a key factor about swollen stems of Jiaobai,It is not sure that cytokinin secreted by itself or U.esculenta P.Henn lead to swelling of gall.Therefore,it is of great significance to reduce the secretory CTK of U.esculenta P.Henn for the purpose of studying the swollen stems of Jiaobai.So we build the pUMA932-CKX-GFP expression vector.It was transformed in U.esculenta P.Henn and the expression of CKX gene was detected by qRT-PCR.The result were as following:1)A variety of bioinformatics softwares were used to obtain 20 D-type cyclins in Jiaobai.According to the phylogenetic tree analysis,it can be concluded that there are 6 subgroups of Jiaobai,which are D1,D5,D6,D7,D2/4,and the only D3 subgroup is absent.The phylogenetic tree branch of Jiaobai and rice was closest,indicating that the relationship between Jiaobai and rice homology was closest.We analyzed the protein of Jiaobai CycD,indicating that the CycD sequence is very highly conserved.2)Through analyzing the results of transcriptome sequencing,we know that Zizla;CycD5;1b are from low to high in the expression analysis of HSD and Zhejiao;Zizla;CycD5;1b can be seen in the difference analysis of the expression in different periods;From the non-inflated to start to enlarge,Zizla;CycD5;1b gene’s expression level was significantly increased,and then the expression level of the lubai was obviously decreased.The expression levels of the male Jiaobai were also relatively low,it was indicated that Zizla;CycD5;1b gene is a key gene during the beginning of swelling.3)Through in situ hybridization and localization experiments,the results showed that the Zizla;CycD5;1b gene has a signal in the apical meristem of Jiaobai,and this gene is expressed in the vascular tissue of Jiaobai during the beginning of swelling,indicating that Zizla;CycD5;1b participates in the cell cycle progression during the expansion.4)The results obtained by using qRT-PCR experiments showed that the expression levels of Zizla;CycD5;1b genes were significantly upregulated in 24 h and 48 h after treatment with t-Zeatin and IAA.The results of the treatment in 24 h were not significant,and it is significantly in 48 h.From this result,it indicates that Zizla;CycD5;1b gene plays a positive regulatory role in the process of Jiaobai expansion by t-Zeatin and IAA treatment.5)By observing of the green fluorescent protein GFP under a fluorescence microscope microscope,we saw the weak green fluorescent protein.The successful expression of the CKX gene was detected in UeT14 and UeT55 by qRT-PCR analysis,and these results indicate that the pUMA932-CKX-GFP cloning vector was successfully transferred into U.esculenta P.Henn.