| Maize is one of the world's three largest food crops,but also an important industrial raw materials,its yield and quality is of great significance to human beings.Among the factors that affect the yield and quality of crops,drought is undoubtedly a very common and influential kind of stress,which is an important problem of agricultural production.Maize is very sensitive to drought stress,the growth and development of Maize under drought stress will be greatly limited,and its yield and quality will be reduced.China is a country with serious water shortage,especially in Northwest China.Crops have been under drought stress during the whole growth period without effective irrigation measures.Therefore,the use of transgenic technology to improve the drought resistance of maize varieties is an effective way to reduce the impact of drought on Maize Yield and quality.Root system is an important organ of plants,the main role is to absorb water and nutrients.The root system of maize is directly related to its ability to resist drought,which affects the formation of the upper parts and final yield of maize.Plant cytokinin and growth are closely linked,the key enzyme in the metabolic pathway is the cytokinin oxidase(CKX),it can adjust the content of cytokinins in plant growth and development,and regulating plant tissue and organs.This research chooses a kind of rice root specific expression of EXP18 D gene promoter and CKX gene linked to transform maize inbred lines,so as to promote the growth and development of root system,in order to obtain drought tolerant maize varieties.In this study,AtCKX3 was introduced into maize inbred lines by pollen tube pathway.Through the screening of T0,T1 and T2 generation of glyphosate herbicides,130 rtransgenic lines were obtained.10 transgenic lines with stronger drought tolerance were selected from T2 generation homozygous lines,The EPSP gene and the target gene CKX3 have been successfully introduced into the genome of C92 inbred line and can be stably inherited and expressed by molecular detection method and some root measurement indicators.The physiological indexes such as plant height,leaf area and 1000 grain weight of the transgenic lines and C92 inbred lines were detected.,the results showed that the transgenic lines were significantly superior to C92 inbred lines.The droplet digital PCR(ddPCR)method to evaluate the exogenous gene copy number in genetically modified crops(GM crops)as the example from genetically modified maize(Zea mays L.)was developed,and some T0 generation transgenic maize plants with single copy of EPSP gene were selected from all the samples to be detected.The results were also compared with those from quantitative real-time PCR(qRT-PCR),as can be seen from the experimental datas,the results from qRT-PCR(TaqMan)and ddPCR for EPSP were consistent,especially the single copy detection results of both in a high degree of consistency.And the ddPCR method is easy to operate,the result is repeatable and accurate.Therefore,the ddPCR will be well developed as a novel method for estimating transgenic copy number with high accuracy,which may be widely used in the exogenous genes copy number analysis in GM crops in the near future. |
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