Enetic Diversity Of Soybean Rhizobia Of Changbai Mountain

rhizobium of soybean is a kind of living microbes, symbiosis with legumes Leguminous plants through photosynthesis manufacturing supply of organic matter to rhizobia;Rhizobium biological nitrogen fixation of nitrogen fixation in leguminous plants, is the highest efficiency.Using biological nitrogen fixation of rhizobium can reduce fertilizer application of soy, enriching the soil fertility, improve soil.The main material of this study is from the soybean plants in changbai mountains different rhizobia, YMA medium through continuous generation of cultivation, separation and purification of soybean rhizobia.To study the genetic diversity of changbai mountains rhizobium of soybean, mainly of rhizobia recA conservative gene, symbiotic nitrogen fixation (nif) plasmid gene genetic study, through agarose gel recovery after PCR sequencing diversity analysis.Through this study the following conclusions:1、through the optimization of YMA medium, choose the suitable for the growth of the changbai mountains Rhizobium of Soybean YMA culture medium is:mannitol10g, yeast extract4g, agar16g, MgSO40.2g, NaCl0.3g, K2HPO40.25g, KH2PO40.25g, adjusted PH=7.1;2、each contained in the root nodules of rhizobium is a kind of rhizobia, through continuous successive transfer culture of rhizobia, YMA medium can be isolation and purification of rhizobia;3、continuous generation of cultivating soybean rhizobia YMA medium on its optimal concentration of diluted for1/2-7;4、-80℃,be stored under the rhizobium fredii, The best suitable to the growth of glycerin concentration75%of glycerol while the rhizobium of soybean storing in the refrigerator of-80℃;5、Microscopically, observing with Gram staining, the rhizobium of soybean length is about1μm negative;6、to the optimized of PCR reaction process is:Initial denaturation95℃for2min.35cycles into the circulatory system, denaturation94℃for1min, annealing temperature63℃for1min, extending72℃for2min, the end of the cycle, The final extending72℃for5mm;7, with nitrogen fixation gene nif constructed phylogenetic tree can be seen, five regions of the changbai mountains of rhizobium of soybean evolved from two directions.C-7,A-5,D-1on a big the branch,can be seen as A generic, yenji, erdao,and helong have the same rhizobium of soybean of the genus. A-2, E-3, C-4, A-2, C-3on a large branch may be regard as A generic, yenji, helong and antu have the same rhizobium of soybean of the genus.D-2, A-4, D-4,E-6,D-6in A large branches can be regard as A generic, yenji, erdao and antu have the same rhizobium of soybean of the genus. B-4, B-1, E-2,D-3,D-1,A-1, E-5, A-5,B-2, B-3on a large branch can be regard as A generic, yenji, longjing, erdao and antu have the same rhizobium of soybean of the genus.longjing area of sample belongs to A genus.E-3and Sinorhizobium sp. On the same branch, may be regarded as the same rhizobium of soybean;8, with recA housekeeping gene constructed phylogenetic tree can be seen, five regions of the changbai mountains of rhizobium of soybean evolved from two directions,E-2, A-3,C-1on a branch, can be seen as A generic, yenji, helong and antu have the same rhizobium of soybean of the genus.C-2, A-5, D-1on a branch, can be seen as A generic, yenji, helonf and erdao have the same rhizobium of soybean of the genus.C-4,E-5,E-3,E-4on a branch, can be seen as a generic,helong and antu have the same rhizobium of soybean of the genus. B-5、A-1、A-4、B-2、 B-4、B-3、B-1、E-1、D-2、D-3gathered in a large branches, yenji, longjing, erdao and antu have the same rhizobium of soybean of the genus. longjing area of sample belongs to A genus.