| Channel catfish(Ictalurus punctatus)is one of the most abundant catfish species in North America,and Omega-3 fatty acids have a positive impact on health.The objectives of this study were to determine the effectiveness of the Masou salmon △5-desaturase transgene driven by a beta-actin promoter for improving Omega-3 fatty acid production in F1 transgenic channel catfish,Ictalurus punctatus together with proximate analysis,as well as to determine pleiotropic effects of disease resistance.Additionally,a novel approach of using multiple transgenes to alter multiple steps in a pathway,which has not been previously reported in aquaculture species was utilized.Muscle samples were obtained from F1 offspring and lipid was extracted.The content of Omega-3 fatty acid was quantitatively determined by gas chromatography-mass spectrometry.The pleiotropic effect of Omega-3 fatty acid was determined by the analysis of muscle components(moisture,ash,fat content,moisture content,protein content).In this study,apo14-masousalmon-elovl2 gene was successfully inserted into chromosome 1 of Channel Catfish by microinjection and RISPR/Cas9-mediated technique.And in addition,the insertion of target genes on chromosomes 2 and 7 of Channel Catfish was also achieved in another Channel catfish.Fatty acid detection and analysis showed that the transformation of △-5 desaturase driven by beta-actin promoter had a positive effect on the production of Omega-3 fatty acid in transgenic catfish.The omega-3 fatty acid production rate of F1 female catfish was24.94%(P=0.026)higher than that of controls,while that of F1 males was 34.87%(P<0.0001)higher than that of controls.The omega-6 fatty acid production rate of F1 female was 6.32% lower than that of control group(P=0.398),while that of male was17.54% lower than that of control group(P=0.001).However,the content of ETA increased by 7%(P = 0.180)in transgenic female catfish,while decreased by 3%(P = 0.270)in transgenic male catfish.As a direct product of △-5 desaturase,Omega-3 fatty acids accounted for 23% of total FMAE in female genetically modified catfish(P=0.038)and30% in male catfish(P<0.05).DPA increased by 16%(P = 0.267)and 27%(P < 0.05)infemale and male catfish respectively,while DHA increased by 32%(P = 0.024)and 47%(P < 0.05)in female and male catfish respectively.There was no significant difference in fat percentage between F1 transgenic female and control group(P=0.350),but the fat percentage of F1 transgenic male was 42.84%lower than that of control group(P<0.001).There was no significant difference in ash percentage between F1 transgenic catfish and control group.There was no significant difference in moisture percentage between F1 female genetically modified catfish and female control group(P=0.911),but the moisture percentage of male genetically modified catfish was higher than that of control group(P=0.002).In addition,the protein percentage of F1 females was 40.23% higher than that of control females(P=0.115).The protein percentage of F1 males was 33.67% higher than that of control males(P<0.001).More over,the average body weight of F1 transgenic catfish was also higher than that of the control group,the disease resistance and hypoxia tolerance of the transgenic catfish were also improved.In this study,the main goal of increasing the proportion of Omega-3 fatty acids has been achieved,but the goal of increasing the total content of Omega-3 fatty acids has not been achieved.In addition to the significant changes in body composition of genetically modified channel catfish,it also shows the effect of pleiotropic effects of transgenic channel catfish,and the transgene appeared to increase uniformity of these traits,but the mechanism is not clear yet.In addition,it was found that the proportion of Omega-3 fatty acids in muscle tissue without skin of channel catfish was higher than that with skin tissue.All these results provides insights for future research. |
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