| 摘要 |
The invention relates to a method for selection in host cells, in which a controlled elimination of a desired DNA sequence occurs and in which the cells in which the above does not occur are sacrificed, by means of the expression of a toxic protein or peptide. The method is characterised in that in step (I), the host cells with (a) the 5' and 3' DNA sequence which is to be subsequently eliminated, flanked by recombinant DNA sequences with a DNA sequence also lying between the recombinant DNA sequences which codes for the toxic protein or peptide and which is controlled by an inducible promoter, are transformed under conditions which lead to the inhibition of the promoter which may be induced and the host cells with (b) a DNA sequence which encodes for a recombinase which recognises the above recombinant DNA sequences and which is under the control of an inducible promoter are transformed under conditions which lead to inhibition of the inducible promoter. The inducible promoters in a) and b) are not the same, or don't have the same inductors. In step (II), elimination of the desired DNA sequence occurs by expression of the recombinase through activation of the inducible promoter (b) and in step (III), the sacrifice of the host cells, in which step (II) has not or not completely taken place, occurs by expression of the toxic protein or peptide through activation of the inducible promoter from (a). In a preferred embodiment the recombinase is a ligand binding domain-recombinase-fusion protein (Rec-LBD). It is further preferred that the desired DNA sequence codes for a marker gene or works as transcription or translation stop sequence. The invention further relates to vectors and host cells containing the sequences (a) and (b), preferably in transgenic plant cells and transgenic plants. |
