Expression Characteristics And Genetic Transformation Of SlSWEET7a In Tomato Fruit

Sugar is an important energy source in the process of plant growth and development.The accumulation and transportation of sugar is very important to the flavor and quality of tomatoes.The transport of sugars in plants is dependent on sugar transporters,which are responsible for the distribution and transport of carbohydrates.SlSWEET7a gene is a member of the new sugar transporter family of SWEETs gene family,floral organs and leaves today for its research mainly concentrated on the model plant,with fleshy fruit of Tomato in this crop,the specific function of SlSWEET7a during the fruit development of research still needs to be.This study uses the method of qRT-PCR to reseach the tomato expression level of SlSWEET7a on SWEETs gene family in different periods and different parts,through the construction of genetic transformation of overexpression and silencing vector were transient expression and tomato fruit,the function of SlSWEET7a gene was validated.The main results are as follows:1)The Solanaceae SWEET7 gene and Arabidopsis SWEETs gene family by phylogenetic analysis found that AtSWEET4?AtSWEET8?StSWEET7a?StSWEET7b and SlSWEET7a had high homology.SlWEET7a as a member of CladeII in SWEETs family,predicts that the sugar in plants as the main carrier of monosaccharide transporters may play a role in plant flowers organ,it also has a role in the regulation of pollen maturation.2)Analysis of qRT-PCR showed that SlSWEET7a gene was specifically expressed in tomato fruit,and the highest expression was in the fruit of green ripe stage.From different parts,green fruit maturity and fruit stalk vascular expression of SlSWEET7a was highest,and the two belong to the phloem tissue,indicating that SlSWEET7a may affect the source organs transport to sink organs facilitated diffusion from sugar.However,the expression of SlSWEET7a was lower than that of control at the time of rotation and red ripening,which indicated that the effect of SlSWEET7a gene was decreased.From flowering to tomato analysis showed that tomatoes 7d after anthesis to mature green stage during this period the expression of SlSWEET7a was up-regulated the expression of fruit maturity,the maturity may on flowers and pollen grains have a regulatory role,just with the bioinformatics analysis results.3)Using the RT-PCR method of the cloned SlSWEET7a gene of tomatoes,and then it was connected with pBI121 vector,constructed the expression vector of pBI121-SlSWEET7a gene;At the same time according to the specific sequence of the gene,the reseach designed primers and gene silencing,amplified by Gateway technique successfully constructed the gene silencing vector pB7GWIWG2(I)-SlSWEET7a.The tomato fruit injection method of Agrobacterium mediated,constructed pBI 121-SlSWEET7a overexpression and silencing vector pB7GWIWG2(I)-SlSWEET7a in the fruit after transient expression,use qRT-PCR method to detect the gene.The results showed that the injection of pBI121-SISWEET7a overexpression carrier of bacteria liquid fruit,the expression of SlSWEET7a gene is more than 7 times of injection of fruit;injection with silent carrier of bacterial fruit than not injected fruit expression decreased nearly 60 times.These results indicated that the overexpression and silencing vector of SlSWEET7a could significantly affect the expression of SISWEET7a,and could be used for further genetic transformation.4)As the tomato Micro-Tom as test materials,of all the constructed vectors for genetic transformation,overexpression lines with kanamycin resistance were screened 34 strains,rooting after PCR were detected by the overexpression of SlSWEET7a 5 strains;silence vectors after infection were screened by herbicide,21 strains were obtained,rooting after culture PCR was detected in 7 strains of transgenic silence,seedling success,further studies are needed to evaluate the specific function.