A NEW LAMBDA RES VECTOR WITH A BUILT-IN TN1721-ENCODED EXCISION SYSTEM

A new lambda replacement vector for construction of genomic libraries was developed which allows the excision of cloned fragments by site-specific recombination from the lambda DNA and conversion into autonomously replicating plasmids. The vector system, derived from lambdaEMBL4, is called lambdaRES. It contains two recognition sites for site-specific recombination from Tn1721 on both sides of the replacement fragment of lambdaEMBL4. Additionally, on one side, there is a plasmid replication origin from Rts1 with a kanamycin-resistance (Km(R)) marker. DNA fragments in the range of 8-14 kb may be inserted between BamHI or Sall sites in the lambda vector. Efficient excision and conversion of plaque-forming units into Km(R) colonies are obtained by infection of Escherichia coli strains harbouring Tn1739tnpR on a F' plasmid. Tn 1739tnpR is a derivative of Tn1721 with a chloramphenicol-resistance-encoding gene (Cm(R)), the lambda cI repressor gene, and a further copy of the resolvase-encoding tnpR gene under control of the tac promoter.