| Rape is an important oil crop in China for a long time. It is a main source of edible oil and protein feeds. How to improve the yield and oil content of rapeseed always is the research focus of rape breeding. Although the main biosynthetic pathways of plant storage material has been studied clearly, but the molecular regulation mechanism of these pathways for external environment and internal factors is not very clear. So, it is necessary that explore the regulation mechanism which these pathways received with a view of gene expression. In this study, we have analyzed promotors of 43 oil and storage protein biosynthesis related genes, detected the expression patterns of main seed transcription factors and storage material biosynthess related genes, researched the effects of Gibberellins (GA) or Abscisic acid (ABA) on storage reserves deposit and agronomical traits of rape and the expression profiles of related genes, and have been screening bZIP transcription factors which maybe response ABA or GA and regulate storage reserves biosynthesis. The results of this study were listed as follows:(1) We have analyzed promotors of 35 oil biosynthesis related key genes and 8 storage protein genes.246 kinds cis element exist in these promotors, including 67 kinds which Frequency of occurrence is more than 70%.15 kinds cis elements are related with plant hormones (such as ABA, GA) cis element,14 kinds are related with the stress,12 kinds are related with light regulation, and 5 kinds are associated with sugar metabolism. These analysis results suggest that plant hormones may play an important regulating role as environmental factors on oil and storage protein synthesis.(2) The expression levels of 6 main seed transcription factors and 20 genes related of storage reserves biosynthesis were determined in different organs of rape variety W6 (Brassica Napus L.) by RT-PCR. Most storage reserves biosynthesis related genes are high express in silique (16/20), then in leaf, bud and flower (14/20), but little in stem and root (3/20); 6 main seed transcription factors mainly express in silique. The expression profiles of these genes during rape seed filling were determined by Q-PCR. The expression profiles of transcription factor genes LEC1, LEC2, WRI, and most storage reserves biosynthesis related genes (15/20) could be described as this: increases first, then decreases until 25 DAF (Days after flowering), and then increases again, but down at the end; transcription factor genes FUS3, ABI3, ABI5 could be classed a steady increase pattern; Storage protein genes Napin and Cruciferin almost no express until 20 DAF, and express a sharply peak at 25 DAF and 30 DAF, respectively. Otherwise, the content of total protein and crude fat in seed was detected during rape seed filling. Total protein and crude fat deposit steady in seed during rapeseed filling, while they have a different deposition patterns with complementary relation at acceleration after 25DAF. The above results show that 25DAF is a key turning point in the development of rapeseed. It may be that the expression changes of genes related storage reserves biosynthesis caused by main seed transcription factors resulting in the different deposition patterns of total protein and crude fat in rapeseed.(3) 100mM GA3 was applied to treat W6 siliques at earlier, middle and later stages (15,25 and 35 DAF) of seed filling. The results are as follows:all 3 GA treatments can increase total sugar and glucosinolates contents, but had not obvious effects on rapeseed oil content; GA applications can decrease protein content in earlier and middle treatments, but increase protein content in later treatment, can slightly decrease seed weight in earlier treatment, and obviously down regulate it in middle or later treatments; Lipid synthesis rate-limiting enzyme gene DGAT was upregulated by GA in earlier treatment, but not affected by GA in middle and later treatments, and this expression pattern was positively related to the change of oil content in rapeseed. The above results show that GA applications during seed filling have no significant effect on rapeseed oil content and may reduced rape yield through decreasing seed weight.(4) 10μM ABA was applied to treat siliques of rape varieties W6 and Xiangzhayou3 at earlier, middle and later stages of seed filling. All 3 ABA treatments can strikingly increase rapeseed weight, oil content, and decrease very long chain fatty acids proportion. ABA applications has different effect on crude protein, total sugar and glucosinolates content in different rapeseed cultivars in all 3 treatments. Otherwise, transcription detection of lipid synthesis related gene show that gene DGAT was upregulated chronically by ABA in all 3 treatments, and it reflect the increase pattern of oil content. The above results show that ABA applications during seed filling can significantly increase weight and oil content of rapeseed and improve rapeseed oil quality.(5) We have screening 3 bZIP genes which high expression in seed and may response ABA or GA by microarray data analysis and Q-PCR detection. These genes were heterologous expression in Arabidopsis. It find that over expression of BnbZIP 12 inhibit growth of plant, over expression of BnbZIP43 could reduce seed setting rate of plant, and over expression of BnbZIP66 could increase seed weight and change seed storage reserves proportion. We guess that BnbZIP12, BnbZIP43, and BnbZIP66 may play an important role in plant vegetative growth, gametes producing or fertilization, regulation of storage reserves biosynthesis, respectively.In conclusion, this study provide a useful reference for the breeding study and actual production of rapeseed, pointed out the direction of further study on the regulation of storage reserves biosynthesis and related transcription factors, bore theoretical and realized meanings to a certain extent. |
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