Isolation And Identification Of Rhizobium From Seabuckthorn And Effects Of Rhizobium On The Growth And Development Of Plants

Seabuckthorn(Hippophae rhamnoides L.),known as vinegar or acid thorn,Hippophae plants of the family Elaeagnacaae,is a plant with a wide range of growth and adaptability.It grows as deciduous shrub or small trees.Sea buckthorn rhizobium is a type of actinomycetes that can co-produce nodules with seabuckthorn.The root of seabuckthorn is well developed and can form a large number of nodules with nodulation actinomycetes,which has high nitrogen fixation ability.Because of the strong adaptability of seabuckthorn ability,and it most grows in desert,plateau,hills and other places relatively poor soil,the growth area of soil has a very significant effect.Some studies have shown that organic matter content of soil is 2.13% and nitrogen content of soil is 0.12% in 6-year-old seabuckthorn forest grown in calcareous soil.Each data are about 1 times higher than ordinary farmland.The results show that the area covered by seabuckthorn can reduce the average annual runoff about 80%,reduce surface water erosion about 75% and reduce wind erosion about 85%.It can be said that the symbiotic system of seabuckthorn and nodulation actinomycetes has great research and development value both in the development of agriculture and forestry and in the ecological protection as well as immeasurable economic value.It is the reason why seabuckthorn has such excellent biological traits that its root can nodulate and fix nitrogen is the key factor.Therefore,the research on rhizobium seabuckthorn is very important to promote the further development and utilization of seabuckthorn.In order to make better use of this plant resource of seabuckthorn and its symbiosis with rhizobium as a microbial resource,it is necessary to fully understand the types of actinomycetes contained in the rhizobium seabuckthron and explore the culture methods of these actinomycetes,included the formation of nodules,the growth and development of plant,as well as nitrogen fixation genes in rhizobia.In this experiment,it was mainly study on the method of isolation and culture of sea buckthorn rhizobia,identified the species of rhizobia,the different types of Seabuckthorn plant growth,and the role of nitrogen fixation and screening of nitrogen fixation genes.The results are as follows:(1)Culture methods for direct isolation of single colonies from seabuckthorn root nodules.Seabuckthorn Rhizobium is an actinomycete belonging to different genera of Actinomycetes and is usually divided into Frankia and non-Franklin's actinomycetes.At present,the isolated and purified non-Frankia actinomycetes are mainly Carnos Actinobacillus(Nocardia)is an actinomycete and Micromonospora actinomycete.The tests showed that the solid medium of Tween-80 was not added during the separation of Actinomyces Frankii and Actinobacillus non-Franklin,and it could be cultured under the constant temperature culture condition of 28°C by the nodule homogenate dilution method.A single colony with obvious morphological differences was formed on the substrate,and the single colonies obtained from the root nodules were separated,and after further separation and purification,pure colonies could be obtained.(2)After repeated isolation and culture,12 single colonies with obvious morphological differences were isolated and purified from seabuckthorn nodule.The colony number was 1-12.After 16 S rRNA sequence analysis,physiological and biochemical analysis,the morphological analysis found 2 Nos.3,4,9 and 11 are Frankia actinomycetes and are designated as strain Fk-17.02,strain Fk-17.03,strain Fk-17.04,strain FK-17.09,strain FK-17.11;1,5,8 No.10 and No.10 strains are actinomycetes belonging to the genus Nocardia and designated as strains QN-17.01,strains QN-17.05,strains QN-17.08,and strains QN-17.10;strains 6,7 and 12 are Micromonospora.(Micromonospora)actinomycetes,designated as strain Mi-17.06,strain Mi-17.07,strain Mi-17.12.(3)Single colony strains of sea buckthorn rhizobia isolated and purified were individually backed and combined back to each other as genus units,and then tested by a single strain as a unit to find that Actinobacillus Frankii could promote the nodulation of seabuckthorn plants directly.Actinomycetes belonging to the genus Cannobacter(Nocardia)can promote plant nodulation.Actinobacteria of Micromonospora and actinomycetes of Nocardia It also has obvious promoting effect on the formation of seabuckthorn root nodule,the growth and development of seabuckthorn plants.It can increase contents ofprotein and chlorophyll in seabuckthorn plant leaves,and the activity of nitrate reductase is enhanced.It also has a certain role in nitrogen fixation system.