Studies On The Physiological And MRNA Differential Display And DNA Methylation During Fleshy Tap Root Development In Radish(Raphanus Sativus L.)

Radish(Raphamus sativus L.)is widely cultivated in the world.. It has a long history,abundant in nutritions and plays an very important role in vegetable production of ournation. Some long-term problems around production restrict the yield and qualityimproving of fleshy tap root, which is the main edible part of radish. The thickening of it'stap root is a very complicated process involving morphology, accumulation of nutritionsand gene expression and so on. Studying on the physiological and gene expression changesand DNA methylation will help us to clarify the mechanism of the thickening in fleshy taproot and ground a good foundation in genetic improving and quality enhancement.In the paper, several physiological changes including carbohydrate cantend andsucrose metabolizing enzyme activities during development of radish fleshy taproot werestudied using high inbred lines of different ratio of root to shoot(R/S). The results showsthat sink activity patterns of taproot were much similar among the four lines. The contentsof carbohydrates including soluble total sugar, sucrose and fructose decrease at thebeginning of taproot enlarging, and to the minimum at the stage of the primary cortex split.The sucrose content of the low R/S line 'NAU-LR-04' was higher than those of three highR/S lines at all stages, while the fructose content was lower in 'NAU-LR-04'. Changes ofactivities of sucrose synthase (SuSy) were similar to sink activities in all the lines, and theactivity of SuSy in low R/S line is higher than those of high R/S lines. It can be inferredthat the SuSy may play an important role in the sucrose level regulation and sink activitycontrol during radish fleshy taproot thickening.The isozyme analysis of leaves and root in radish of 'NAU-LHZ-03'showed that boththe number and intensity of the bands of SOD and EST isozymes in leaves were much highthan in root. For SOD isozyme there was a single band (RI 0.15) that only existed in rootsbut not in leaves, so it can be suggested that this band may have some relationship with thethickening of the fleshy tap root. While for amylase isozyme pattern there were twoidentical bands existed during all the stages of development in both leaves and root, so itcan be inferred that there is no relationship between the thickening of fleshy tap root in radish and amylase isozyme.Differential expressed genes in course of fleshy tap root formation and thickening inradish of 'NAU-YH-05' were studied using mRNA differential display PCR in use of 60pairs of primer. Some differential expressed segments were obtained and cloned, theirsequences were compared with in identity of Blastn entry into GENEBANK. Comparabilityof some segments reached about 86% to 90% including Arabidopsis thaliana CIPK7,Arabidopsis thaliana At3g23000 gene,Arabidopsis thaliana SNF1 related protein kinase(At3g23000; MXC7.3) mRNA,Cucumis sativus RBL1 mRNA, complete cds,Ranunculusmacranthus ribosomal protein L14 (rp114) gene, partial cds,Daucus carota chromasomecomplete genome and so on. There are also some other segments had a low identity andconsidered as unknown genes.MSAP patterns with 21 pairs of primer were studied in leaves and root of radish withdifferent ratio of root to shoot. Results showed that MSAP levels had a trend of increase inroot and the level of MSAP in time D₁ and D₂ of 'NAU-LHZ-03' with a low ratio of rootto shoot were high than that in 'NAURa-DY02', but it was opposite in time D₃. Studying ofMethylation in leaves and root with different stages showed that the full methylation ofgenome DNA were always higher than hemi-methylation of cytosine, it indicated that thefull methylation other than himi-methylation of cytosine took a predominate role in DNAmethylation in different stages and different part of radish.