Plant-induced Inter-genus Horizontal Gene Transfer Of Symbiosis Island Of Azorhizobium Caulinodans Broadens Rhizobial Host-range Specificity

Rhizobium can form nodules with legume and reduce N2 in the air to nitrogen for the plant,at the same time the plant provides carbon and energy source and other nutritional substances for the rhizobium.Up to now,most of the plants only form nodules in root,only a few plants can form nodules in stem.The studied strain Azorhizobium caulinodans ORS571 can form both root and stem nodules on Sesbania rostrata,the genome of ORS571 was sequenced in 2008,a symbiosis island about 87.6 kb was detected in the genome,a lot of transfer gene are in the island,the area also contains three genetic loci associated with symbiosis involves the synthesis and secretion of Nod factors.Current research on horizontal gene transfer were focused on pathogenic bacteria,not much about the rhizobia symbiosis gene,we study symbiosis island in the ORS571 in our experiment.First,we insert kanamycin in the noncoding region of the symbiosis island named Azcl as the doner,receptors are lab strains,including Rhizobium,Mesorhizobium,Sinorhizobium,Bradyrhizobium.The results show that in the experimental range,all of the Mesorhizobium could accept the symbiosis island,Rhizobium and Bradyrhizobium can not,S.medicae can while S.meliloti can not,the frequency is between 10-7 to 10-8.In order to test the transfer frequency with plants,testing the effect to transfer frequency with different host plants(S.rostrata,A.sinicus,Zea mays).The results found that transfer frequency was improved at least 10 times with plants,and transfer frequency was different with different plant.The symbiosis island contain nodulation genes related to the synthesis and secretion of Nod factors,so we detect if the physiological function is changed after receiving symbiosis island in receptor rhizobium,The results show that receptor rhizobium could nodule with a new plant and the nodule have nitrogenase activity.Further to knock out the nodulation genes of the island,we found that when nodD was destroyed in Azc1,it could not nodule with S.rostrata any more,but the conjugant could nodule with S.rostrata.In summary,in our experiment the symbiosis island can be transferred to different genus with the transfer frequency were about 10-7 by means of conjugation,and the transfer frequency can be improved with plant.Then the receptor bacterium acquire the ability to nodule with new host after accept the symbiosis island.