| With the deepening of the national "Rural Revitalization" strategy and the advocacy of "two reductions and one increase",Rhizobium fertilizer,which can provide a large amount of nitrogen nutrition for specific crops,is the hot direction of agricultural fertilizer development in China in the future.The research on the molecular mechanism of "symbiotic nitrogen fixation" of rhizobia can provide a solid theoretical basis for the wider and efficient application of rhizobia.In this paper,Mesorhizobium huakuii 7653 R and Sinorhizobium meliloti 1021 were used as research materials to study the function of GTP binding protein gene typa in symbiotic nitrogen fixation.The results are as follows:typA deletion mutants of Mesorhizobium huakuii 7653 R and Sinorhizobium meliloti1021 were constructed and screened respectively.Their symbiotic phenotypes were completely consistent after inoculating plants,and they all showed the nitrogen fixation defect phenotype of white nodules.Supplementing the mutant strain can restore the symbiotic nitrogen fixation ability.The growth curve showed that the deletion of typA did not affect the growth rate of the strain.The detection of autogenic phenotype showed that the resistance to stress decreased after typA deletion.The competitive nodulation experiment showed that the strain with typa deletion of 7653 R had stronger competitive nodulation ability.Through bacterial double hybrid experiment,it was confirmed that there was interaction between 7653 R typA whole protein and about 150 amino acids at the C-terminal of bacA protein.Further pull-down detection showed that they also had interaction in vitro,and the interaction required a complete high-level structure of the protein(the denatured protein could not prove the interaction by far Western blot).The typA and bacA double gene deletion mutants of Mesorhizobium huakuii 7653 R and Sinorhizobium meliloti 1021 were constructed respectively.There was no significant difference between them and the single gene deletion mutants after inoculation.They all showed nodulation and non nitrogen fixation,but the number and weight of nodules were significantly different from their single gene mutants.Autogenetic phenotype test showed that the deletion of typA and bacA would significantly affect the stress resistance of the strain.Quantitative detection showed that the effects of typA and bacA deletion on the stress resistance of the strain were roughly the same,and the stress resistance of typA and bacA double deletion mutants was not significantly different from that of single gene deletion strains,indicating that the effects of typa and Baca on the stress resistance of the strain were the same,and there was no superposition effect.Among the strains forming amorphous nodules,typA is the key gene for symbiotic nitrogen fixation between rhizobia and legumes,and its effect on stress resistance and symbiotic phenotype is similar to bacA gene;The study of typA gene can better analyze the differentiation mechanism of rhizobia and provide a theoretical basis for the research and development of new engineering rhizobia fertilizers... |
