Cloning And Expression Analysis Of Parthenocarpy Related Genes CsWDR36 In Cucumber

Cucumber(Cucumis sativus L.)is one of the widely cultivated vegetable crops in the world,has an important role in consumption and production.The area of protected cultivation is very large in our country,low temperature,high humidity and male flowers indispensable factors always appeared in the protected cultivation,they would led to poor pollination and fertilization,and the resulting in the decrease of yield and quality of cucumber.That is the most important problems to be solved the cucumber production in our country now.Parthenocarpy enables fruit set in absence of the pollination/fertilization,which could ensure the yield,and enhance its commercial quality.However,the inheritance and regulation mechanism of parthenocarpy are still unclear.New resources relatived with parthenocarpy collecting and studying can make a sense to improve cucumber cultivars.In this study,the parthenocarpic lines 'EC1' and non-parthenocarpic lines '8419s-1' as test materials to study the function of candidate genes.Basing on the QTL mapping of cucumber parthenocarpy,we found the genes related with fruit develpoment by bioinformatics analysis,RNA-Seq technology and re-sequencing,then we predicted the candidate genes related related to parthenocarpy.The CsWDR36 was choosed to be cloned and for the analyses of expressional level in early fruit development,in different organs,and their response to different hormones.We also studie the subcellular location of CsWDR36 protein.Finally,genome-wide characterization of WDR genes family in cucumber enable us to understand the networks of WDR proteins.The main results were as following:1 Prediction and expression analysis of parthenocarpy candidate genes in cucumber.In the study,there were 98 genes in the localization of the major parthenocarpy QTL,35 genes were identified referred to biological process e.g.cell cycle,cell growth,hormone signal transduction.Co-analyzing with the transcriptome data,6 different expression genes were selected and their potential function were annotated by KEGG.The expression patterns of the 6 genes responding to differently developed cucumber fruits were analyzed by real-time RT-PCR.Only the CsCRT1 and CsCOL3 have up-regulate expressed during parthenocarpy fruit set;the CsWDR36 expressed contrary between the abortion and pollination progress.So we predicted CsCRT1,CsCOL3 and CsWDR36 were the candidate genes.With resequencing of 'EC1' and '8419s-1',we found there were amount of SNPs in the CsWDR36 and CsSTK.Therefore,CsWDR36 were regarded as the most candidate genes and used in the further study.2 Genome-wide association analysis of WDR gene families in cucumber.The characteristic eukaryotes domain sequence of WDR was generated by Hidden Markov Model(HMM)profile(PF00400),which were employed as a query to search against the cucumber genome database,142 potential WDR genes were identified in the latest cucumber genome.All the CsWDR genes were mapped onto chromosomes 1-7,the WDR number on each chromosomes was 28?16?32?19?16?19 and 12 respectively.According to the structure of these genes,they were divided into 12 groups,and only containing WD domain accounted for about 70%of the total.Phylogenetic analysis categorized the WDR proteins into seven subgroups.We predict the function of the group ? may be associated with the development of embryo and gametogenesis;Group ? gene may be involved in cell division process;Group ? may play a role in salt stress response mechanism and ABA signal pateway;The first group ? genes may regulate the growth and development of plant;?a set of genes may be associated with anthocyanin synthesis;The ?b group may be associated with drought stress response mechanism.By the way,CsWDR36 were classed into the ? group,which could parcipatied in salt stress response mechanism and ABA signal pateway.By analyzing the function of the gene annotation in cucumber WDR gene families involved in many important biological pathways and mostly concentrated in the cell.3 Cloning and Expression Analysis of the parthenocarpy Gene CsWDR36 in Cucumber.In present study,the parthenocarpic lines 'EC1' and non-parthenocarpic lines'8419s-1' were as test materials to clone the cDNA of CsWDR36,there were two different transcription of CsWDR36,named as CsWDR36-1 and CsWDR36-2,and the full length of them were 2676 bp and 1038 bp respectively.We found there was no obvious difference in the two transcription on the hydrophilicity,signal peptide and chemical properties,expect the structure.Similarity analysis showed the sequence of CsWDR36 protein compared with other sequences of WDR in garbanzo beans,rape and melon shared 70%-80%in homology.The phylogenetic tree analysis showed that CsWDR36 was clusterd together with melon,the similarlity values was 82.15%.According to the results of quantitative and semi-quantitative expression,we found that the expression of CsWDR36-1 and CsWDR36-2 were higer than in other organs,and was not sensitive to hormone treatment.The expression trend of CsWDR36-l and CsWDR36-2 was almost unanimously in 'EC 1' and '8419s-1',the expression level was very low before blossoming,at 1dpa to their peakand then decline.But the expression level of CsWDR36-2 was higer than CsWDR36-1 in the whole early development stage.Therefore,CsWDR36-1 and CsWDR36-2 could be involved in the parthenocarpy process in the cucumber.