Regulation Of Oil Content By Interference Expression Of The Arabidopsis PPC1 Gene In Rapeseed (Brassica Napus L.)

Rapeseed (Brassica napus L.) is the world's second-biggest oil crop, and rapeseed oil, close to soybean oil and palm oil of the front two positions, is the third-largest edible oil that circulated in the international market. Our country is the world's largest producer of rape, and the planting area and output of it rank first in the world. Increasing seed oil content of rape is always the important goal for rape breeders, but it is very slow through conventional breeding to increase oil content. So, with the rapid development of genetic engineering techniques, improving rapeseed oil content through transgenic methods gets into good graces by more and more people.Phosphoenolpyruvate carboxylase (PEPC) is considered to be a key enzyme in controlling rapeseed protein/fat ratio. It can regulate the flowing direction of PEP which is the common substrate of the biosynthesis of fat and protein through inhibiting PEPC genes to control target trait, so it can regulates protein/fat ratio from the level of gene expression and biochemical metabolism. Rape and Arabidopsis thaliana are both plants of cruciferae brassica and their genomic homologies are very high. Therefore, Arabidopsis thaliana gene can be directly used in rape. This research around Arabidopsis thaliana PPC1 carried out the following job:1. Cloned Arabidopsis thaliana gene PPC1 and constructed RNAi vector using plasmid pFGC-5941:In this study, designed a pair of primers according to the sequence of Arabidopsis thaliana gene PPC1, amplified Arabidopsis thaliana PPC1 gene fragment by PCR, then cloned into pMD18-T Vector. After confirmed by DNA sequence, the gene was connected to plasmid pFGC-5941 in the form of forward and reverse to construct RNAi expression vector,and then the 35S promoter should be replaced by Napin promoter. 2.86lines To transformation plants was obtained through Agrobacterium-mediated method.There are only 41 lines survived, when these lines which have been transplanted into flowerpot in greenhouse over a period of time were transplanted into field. Through PCR detection, Southern hybridization and insertional locus boundary sequence analysis, We got 21 positive independent transgenic lines3. According to the genetic analysis of some T1 generation, the target gene was mostly inserted by unit point and trebl points,but there some lines belong to multi-loci integration.4. The quality of analysis indicated that the oil content of some To transformed lines were higher than the untransformed ones to some extent, while the other oil content is similar as the control. For fatty acids content,there was no significant change in To transgenic plants,the result shows that the interference expression of Arabidopsis thaliana PPC1 gene has no effect on the biosynthesis of fatty acids.In a word, this research cloned Arabidopsis thaliana PPC1 gene fragment, constructed the RNAi expression vector and translated rape lines of CY2. After a series of molecular testing,got some positive independent transgenic plants,th en analysis of their genetic regularity and quality.