Identification And Evaluation Of Insect-resistant Transformants Of Soybean BT Gene

Among the main food crops in our country,soybean contains a variety of amino acids and vitamins,rich in nutrition and easy to grow,has become an indispensable and important food material.But in recent years,soybean pest has caused a great impact on soybean production in our country,and the usual chemical control technology not only increases the cost,but also causes environmental pollution,and also poses a threat to the food safety of soybean.The breeding of insect-resistant soybean varieties with high yield by plant genetic engineering has gradually become an effective solution.In current genetic engineering techniques,the BT?bacillus thuringiensis,BT?gene was first applied to crop breeding and has shown good performance in insect resistance.By studying transgenic technology,when the BT gene is introduced into crops,it can fully show good insect resistance.In this test,180 strains of the insect-resistant gene Cry1Iem,120 strains of the insect-resistant gene Cry8-like,and 90 strains of the insect-resistant gene Cry1AB13-2 were taken as the research objects.By using conventional PCR detection analysis,Southern hybridization,and fluorescent quantitative PCR technology to conduct molecular identification of the insect resistance gene-transformed strains,and then carry out indoor extermination of these three insect resistance gene-positive plants to select agronomy Traits to identify the insect resistance functions of different insect resistance genes.The main results of this experiment are as follows:1. The T₆strain transformed with the insect resistance gene Cry1Iem soybean,the T₅strain transformed with the Cry8-like soybean,and the T₄strain transformed with the insect resistance gene Cry1AB13-2 were selected as test materials,and 10 of them were selected for routine PCR detection.The target genes Cry1Iem,Cry8-like,Cry1AB13-2,Bar,and promoter were detected in the T₆strain of the insect-resistant gene Cry1Iem soybean,the T₅strain of Cry8-like soybean,and the T₄strain of Cry1AB13-2 soybean.Ca MV35S,Nos target segment.The results showed that all the target genes in the soybean plant"Ji Nong 28"were successfully introduced and integrated in a single copy,and were stably inherited.2. Using the Bar gene as a probe,five lines were selected from the T₆strain transformed with the insect resistance gene Cry1Iem soybean,the T₅strain transformed with Cry8-like soybean,and the T₄strain transformed with Cry1AB13-2 soybean for Southern hybridization.The results showed that the target genes were integrated into the soybean genome of"Ji Nong 28"in a single copy.3. The results of fluorescent quantitative PCR showed that the relative expression levels of target genes in leaves and stems of the 6 plants transformed with Cry1Iem insect resistance genes were:5.46,5.89,6.55,4.61,4.22,3.97;1.16,1.22,1.50,0.88,0.78,1.02.The relative expression levels of 5 transgenic Cry8-like insect-resistant genes at plant roots were selected as:6.28,2.43,10.52,2.45,1.37.The relative expressions of 5 transgenic Cry1AB13-2 insect resistance genes in the root,stem and leaf of the plant were:1.25,1.79,1.98,2.53,1.55;3.04,3.78,2.82,3.97,2.45;6.11,6.67,5.89,7.12,5.32.No fluorescence signal was detected in the non-transformed receptor"Ji Nong 28"soybean plants.The expression of the same part of different strains was different,and the expression of different parts of the same strain was different.4. The identification results of outdoor anti-carnivorous insects show that:The insect-eating rate of the"Jinong 28"soybean recipient plants was 6.97%,and the insect-eating rate of the Cry1Iem transgenic plants was 3.41%,3.56%lower than that of the untransformed plants.The worm-eating rate of the gene lines was 3.14%,which was 3.83%lower than that of the untransformed plants,and the resistance level reached the resistance level;The resistance level also reaches the resistance level.At the same time,there was no significant difference in the other traits between the insect-resistant gene Cry1Iem and the other two insect-resistant gene lines.This transgenic insect-resistant gene line is superior.5. The results of indoor anti-sticky insect identification showed:Within 72 hours,the body weight of the armyworms feeding on the leaves of the insect-resistant genes Cry1Iem,Cry8-like,and Cry1AB13-2 were significantly lower than those of the armyworm eating the leaves of the soybean plants.The susceptibility index of the soybean variety"Jinong 28"reached 72.5397±1.2993,which belongs to the susceptibility level,and the insect resistance levels of the Cry1Iem gene plants,Cry8-like gene plants and Cry1AB13-2 gene plants are MR-Above medium resistance?40?IF<50?.This indicates that the soybean lines transformed with the insect resistance genes Cry1Iem,Cry8-like,and Cry1AB13-2 have significant insect resistance,while the soybean plants transformed with the Cry1AB13-2 insect resistance gene perform better in insect resistance.The susceptibility index of the transformed strain Cry1AB13-2-22 reached15.3703±0.6677.