RT Journal Article SR Electronic(1) A1 Van Mellaert, Lieve A1 Mei, Lijuan A1 Lammertyn, Elke A1 Schacht, Sabine A1 Ann, JozefYR 1998 T1 Site-specific integration of bacteriophage VWB genome into Streptomyces venezuelae and construction of a VWB-based integrative vector JF Microbiology, VO 144 IS 12 SP 3351 OP 3358 DO https://doi.org/10.1099/00221287-144-12-3351 PB Microbiology Society, SN 1465-2080, AB Summary: The temperate bacteriophage VWB integrates into the chromosome of Streptomyces venezuelae ETH14630 via site-specific integration. Following recombination of the VWB attP region with the chromosomal attB sequence, the host-phage junctions attL and attR are formed. Nucleotide sequence analysis of attP, attB, attL and attR revealed a 45 bp common core sequence. In attB this 45 bp sequence consists of the 3' end of a putative tRNAArg(AGG) gene with a 3'-terminal CCA sequence which is typical for prokaryotic tRNAs. Phage DNA integration restores the putative tRNAArg(AGG) gene in attL. However, following recombination the CCA sequence is missing as is the case for most Streptomyces tRNA genes described so far. Adjacent to VWB attP, an ORF encoding a 427 aa protein was detected. The C-terminal region of this protein shows high similarity to the conserved C-terminal domain of site-specific recombinases belonging to the integrase family. To prove the functionality of this putative integrase gene (int), an integrative vector pKT02 was constructed. This vector consists of a 23 kb HindIII-Sphl restriction fragment of VWB DNA containing attP and int cloned in a non-replicative Escherichia coli vector carrying a thiostrepton-resistance (tsr) gene. Integration of pKT02 was obtained after transformation of Streptomyces venezuelae ETH14630 and Streptomyces lividans TK24 protoplasts. This vector will thus be useful for a number of additional Streptomyces species in which a suitable tRNA gene can be functional as integration site., UL https://www.microbiologyresearch.org/content/journal/micro/10.1099/00221287-144-12-3351