Study On Stress Resistance In Recombinant Human Lysozyme-overexpression Maize

Maize is an important food,feed,and economic crops which plays an important role in national food security.However,the factors of pests and diseases have been seriously affecting the output and quality in maize.The traditional genetic improvement methods have the problem of long period and poor resources.Using genetic engineering technology for maize genetic improvement is an important way to solve the above problems in corn production.With the development of molecular biology and the identification and functional verification of new genetic resources,an efficient maize genetic transformation methods are becoming more and more important in the improvement of maize stress resistance traits.Recombinant human lysozyme(rh LZ)gene is a protein stimulating factor that causes the gram-positive bacteria cell wall rupture under osmotic pressure and bacteriolysis,it can also resist fungal infection which contain chitin in the cell wall and inactivate viruses that invade the body by reacts with the negatively charged viral proteins.The successful example for the application of lysozyme in plants is that the rice transformed into the animal lysozyme gene obtain transgenic plants resistant to rice blast.On this basis,the system of maize immature embryo transformation was optimized,the vector construction of rh LYZ,the acquisition and stress resistance analysis of transformed rh LYZ maize.The main findings are as follows:1)The transformation efficiency was stabilised at 30% eventually by increasing the induction rate of embryogenic callus through adding low concentration plant growth regulator NAA as a cofactor,increasing the regeneration rate of embryogenic callus through adding low concentration of Na Cl and TDZ and increasing the infection of immature embryo through adding a certain concentration of Mg Cl2.2)The target gene rh LYZ was separately constructed with the expression vectors p BI121 and p CAMBIA1300 through genetic engineering and transformed into corn.The constructed recombinant plasmid was successfully transformed in maize and the transformed plant of rh LYZ corn T0 was obtained.The successfully transformed rh LYZ maize T0 plants were generated rh LYZ homozygotes by self-crossing two successive generations.3)The biotic stress of homozygotes through Botrytis Cinerea and Fusarium Verticillioids.The results showed that the transgenic rh LYZ maizes had very significant inhibitory effect to the Botrytis Cinerea and Fusarium Verticillioids and showed resistance to corn root rot,stem rot,and ear rot,as well as broad-spectrum fungal resistance such as seed and cob resistance.4)The abiotic stresses of homozygous under high temperature,drought,extreme p H,salt and heavy metals showed that the transgenic rh LYZ maizes had good resistance to drought,salt and heavy metals.The above research results optimized the transformation system of maize immature embryos,verified the good stress resistance of rh LYZ in maize,and provided a new technical system and germplasm resources for maize resistance breeding.