Preliminary Studies On The Impact Of Cottonseeds Oil Quality By Simultaneous Silencing FAD2-1and FatB Genes

Cotton is one of the most important fiber plants, and six major oil crops. However, relatively highsaturated fatty acid contents in cottonseed oil (containing24%palmitic acid C16:0), which increasedhuman cardiovascular risk; the Easy oxidation stability of the polyunsaturated fatty acids is relatively high,reach to55%; But both stability and good for the health of human body fatty acid, oleic acid (C18:1)content is low. How to increase the content of the oleic acid in cottonseed oil effectively and improvecottonseed oil nutritional value, which is beneficial to people’s health, is an important problem to be solved.The main cultivated varieties upland cotton is allotetraploid, polyploidy genetic material strictly limitthe mutation breeding technology and the using of traditional knockout technology; Using fatty acidmetabolism of gene engineering to manipulate fatty acid metabolism in cotton, targeted improvements andquality of cottonseed oil component, has the important value in producing. Out of many geneticengineering methods, the RNA interference (RNAi) technology of high efficiency, specificity, and thesuperiority on the study of gene families and polyploid, will have broad prospect of application in cottongenetic improvement of quality.With cottonseed oil as research object, from the perspective of fatty acid metabolism, we use RNAitechnology to change the material flow direction of fatty acid metabolism, by synchronizing inhibiting lipidsynthesis two key enzyme gene expression level, reducing the easy oxidation of saturated fatty acid andinstability of polyunsaturated fatty acid synthesis in cottonseed oil, improve cotton in both stable andhealthy oleic acid content, achieve the goal of improving cottonseed oil nutritional value.1. Application of bioinformatics software to analysis and comparison of the two target genes functionregional sequence of FAD2-1and FatB in cotton which have reported. Respectively amplification obtainpartial function of FAD2-1and FatB fragments (426bp and501bp respectively) on the foundation of ourprevious studies. while cloned cotton specificity SP promoter, by introducing the specific primers ofenzyme loci, with the method of combining recombinant technology and Gateway technology, werespectively construct the double genes genotype interference expression vector p35HK-FAD2-FatB andseed specificity interference expression vector pBISP-FAD2-FatB.2. Through the leaf discs method into tobacco, obtained the resistance of tobacco by Kana screening.extract genomic DNA for PCR verification, we get positive transgenic tobacco plants. Formed later periodtotal RNA extraction of tobacco seeds, reverse transcription PCR for cDNA and validation, identification atthe RNA level of double genes expression. Through gas chromatography determination of geneticallymodified and contrast tobacco fatty acid type and content of each component, the results show that: tobaccoseeds mainly contained palmitic acid, stearic acid, oleic acid and linoleic acid four kinds of fatty acids,compared with contrast tobaccos, transgenic tobacco seeds of palmitic acid and stearic acid havesignificantly lower, linoleic acid also has significantly reduced, oleic acid content is increased significantly. 3. Through the method of genetic transformation by agrobacterium infect cotton hypocotyl to cottonYZ-1and Xinluzao33, with transfer culturing and Kana screening,70resistant plant were abtained,extraction resistance of cotton genomic DNA for PCR verification, abtained11positive transgenic cottonplants.