Rhizobium vector system

Details Rhizobium vector system Rhizobium vector system

C - Chemistry, Metallurgy

12

N

150/15.2, 195/1.

C12N 15/00 (2006.01) C07K 14/195 (2006.01) C12N 15/64 (2006.01) C12N 15/74 (2006.01) C12N 15/90 (2006.01) C12Q 1/02 (2006.01) C12Q 1/34 (2006.01) C12Q 1/68 (2006.01)

Patent

CA 1236037

ABSTRACT OF THE DISCLOSURE Symbiotic genes of Rhizobium bacteria are isolated and identified, for cloning into other cells etc., by a process which involves introducing into the chromosomal DNA of Rhizobium a promoterless lac operon from E. coli. This is accomplished by use of a novel construct of the promoterless lac operon, a transconjugant such as defective mu-d I (kan, lac) phage and a suitable suicide vector such as pGS6. The transconjugant Rhizobia so formed is selected for kanR, to isolate cells which have taken up the lac operon. On a random basis, some of the transconjugants have inserted the lac operon appropriately to a sym gene promoter which now acts as promoter for the lac operon. Now, in the appropriate stimulative environment for the sym gene promoter, the lac Z gene transcription to produce .beta.-galactosidase is initiated instead of the usual sym gene function. The generation of .beta.-galactosidase is readily detected and analysed, so that cells in which the lac operon fraction has entered the sym gene can be identified, cultivated, and their genome DNA analysed to locate the gene.

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