Abstract: The present invention pertains to a novel method for the generation of a vector construct suitable for gene editing applications which comprises a fixed pair of predetermined expressible guide RNA (gRNA) sequences. The method of the invention allows for an easy construction of such vectors and provides in addition thereto vector libraries for the expression of fixed pairs of gRNAs. The vectors of the invention may be advantageously used to cut out larger genomic DNA sequences, or alternatively, to simultaneously introduce mutations in the genome without a loss or larger genomic sequences. Hence, the system of the invention provides for many molecular genetic approaches for genome alteration.