Resistance Of Transgenic Bt Rice And Maize To Spodoptera Frugiperda

Spodoptera frugiperda(J.E.Smith),which originates from tropical and subtropical regions of the Americas,is rated as a global migratory agricultural pest.The insect infested the country via Myanmar in January 2019 and is currently in a sustained outbreak phase,which has resulted in serious damage to food and cash crop yields such as maize and rice.The disadvantages of using chemical methods to control pests are increasing.Transferring Bt insect-resistant gene into main food crops can be used as an effective way to control fall armyworm.Bacillus thuringiensis can produce insecticidal crystal proteins that are only effective against target pests,with high insecticidal activity and specificity.By using genetic engineering technology,it is introduced into crop varieties to cultivate transgenic crops with insect resistance,which can effectively control pests without causing environmental pollution and human safety.In this study,Cry53 A,Cry56Aa1 and Vip3Aa29 were used as research objects.After codon optimization,Cry53 A + Vip3Aa29,Cry56Aa1 + Vip3Aa29 were transferred into Shuhui 818(R818)to obtain transgenic rice,and Cry53 A was transferred into KN5585 to obtain transgenic maize.All transformed plants obtained were subjected to a series of molecular level tests as well as identification and screening for insect resistance.1.S.frugiperda collected in Sichuan Province were identified as ’ corn strain ’ by COI and Tpi gene identification methods.After three generations of continuous feeding on rice,a small amount of ’ rice strain ’ was detected in each generation of larval samples.2.The results of Southern blot showed that 16 lines of transgenic Cry53 A + Vip3Aa29 rice were single copy and 5 double copy lines.In transgenic Cry56Aa1 + Vip3Aa29 rice,16 lines were single copy and 4 lines were double copy.There were 49 lines of Cry53 A transgenic maize with single copy,7 lines with double copies,and 4 lines with three copies.3.The q RT-PCR analysis showed that Cry53 A,Cry56Aa1 and Vip3Aa29 could be normally transcribed in different tissues of single-copy transgenic rice,and the relative expression levels of exogenous genes at filling stage were leaf > stem > panicle.Cry53 A can also be normally transcribed in different tissues of single-copy transgenic maize.The relative expression level at tasseling stage is leaf > stem > bract > silk > female ear > tassel.The transcription level is abnormal in double-copy rice and maize lines,showing a decrease in relative expression.4.The results of Elisa showed that the expression of Bt protein in single-copy transgenic rice and maize was significantly higher than that in double-copy plants at different stages.In transgenic rice,protein content was higher in both leaves and stalks at the tassel stage and significantly higher than in young ears.Cry53 A protein expression ranged from 1.861-11.990 μg/g(FW)and Cry56Aa1 protein expression tended to range from 2.060-13.316 μg/g(FW).In transgenic maize,the expression of Cry53 A protein was higher in both leaves and stems during the filling stage of maize.The highest expression level in leaves was between 7.665-10.500 μg/g(FW),and the protein content in grains was the lowest,tending to be between 1.262-1.542 μg/g(FW).5.Bioassay results showed that Cry53 A + Vip3Aa29 and Cry56Aa1 + Vip3Aa29 transgenic rice had obvious insecticidal activity against S.frugiperda,but the effect was different among individual plants.Cry53 A transgenic maize had obvious toxicity and growth inhibition to S.frugiperda.The double-copy strain had low inhibitory effect on the growth of S.frugiperda.