Biomaterial Database

Plasmid pIJ699, a multi-copy positive-selection vector for Streptomyces. 1988

T Kieser, and R E Melton

John Innes Institute, Norwich, U.K.

A plasmid vector, pIJ699, which provides positive selection for cloned DNA, was constructed using the replication functions of the Streptomyces wide-host-range multi-copy plasmid pIJ101. The selection for inserts is based on the principle that plasmids with long uninterrupted perfect palindromes (inverted repeats) are 'not viable' in bacteria. For cloning, pIJ699 is digested with BglII. This produces two fragments, one of which is the linearized vector, with two arms of the palindrome at its ends, and the other is a 'spacer' which is needed to keep the inverted repeat sequences apart. The vector fragment is separated from the 'spacer' fragment and ligated with the DNA to be cloned. Plasmids with a fragment of cloned DNA, but not the circularized vector, give rise to thiostrepton-resistant transformants in Streptomyces lividans. The inverted repeat sequences contain a strong transcription terminator which reduces transcriptional read-through both in and out of the cloned fragment. This improves the stability of many hybrid plasmids and facilitates the study of the regulation of cloned genes.

UI	MeSH Term	Description	Entries
D009710	Nucleotide Mapping	Two-dimensional separation and analysis of nucleotides.	Fingerprints, Nucleotide,Fingerprint, Nucleotide,Mapping, Nucleotide,Mappings, Nucleotide,Nucleotide Fingerprint,Nucleotide Fingerprints,Nucleotide Mappings
D010957	Plasmids	Extrachromosomal, usually CIRCULAR DNA molecules that are self-replicating and transferable from one organism to another. They are found in a variety of bacterial, archaeal, fungal, algal, and plant species. They are used in GENETIC ENGINEERING as CLONING VECTORS.	Episomes,Episome,Plasmid
D003001	Cloning, Molecular	The insertion of recombinant DNA molecules from prokaryotic and/or eukaryotic sources into a replicating vehicle, such as a plasmid or virus vector, and the introduction of the resultant hybrid molecules into recipient cells without altering the viability of those cells.	Molecular Cloning
D004262	DNA Restriction Enzymes	Enzymes that are part of the restriction-modification systems. They catalyze the endonucleolytic cleavage of DNA sequences which lack the species-specific methylation pattern in the host cell's DNA. Cleavage yields random or specific double-stranded fragments with terminal 5'-phosphates. The function of restriction enzymes is to destroy any foreign DNA that invades the host cell. Most have been studied in bacterial systems, but a few have been found in eukaryotic organisms. They are also used as tools for the systematic dissection and mapping of chromosomes, in the determination of base sequences of DNAs, and have made it possible to splice and recombine genes from one organism into the genome of another. EC 3.21.1.	Restriction Endonucleases,DNA Restriction Enzyme,Restriction Endonuclease,Endonuclease, Restriction,Endonucleases, Restriction,Enzymes, DNA Restriction,Restriction Enzyme, DNA,Restriction Enzymes, DNA
D005822	Genetic Vectors	DNA molecules capable of autonomous replication within a host cell and into which other DNA sequences can be inserted and thus amplified. Many are derived from PLASMIDS; BACTERIOPHAGES; or VIRUSES. They are used for transporting foreign genes into recipient cells. Genetic vectors possess a functional replicator site and contain GENETIC MARKERS to facilitate their selective recognition.	Cloning Vectors,Shuttle Vectors,Vectors, Genetic,Cloning Vector,Genetic Vector,Shuttle Vector,Vector, Cloning,Vector, Genetic,Vector, Shuttle,Vectors, Cloning,Vectors, Shuttle
D013302	Streptomyces	A genus of bacteria that form a nonfragmented aerial mycelium. Many species have been identified with some being pathogenic. This genus is responsible for producing a majority of the ANTI-BACTERIAL AGENTS of practical value.
D013728	Terminator Regions, Genetic	DNA sequences recognized as signals to end GENETIC TRANSCRIPTION.	Terminator Sequence,Transcriptional Terminator Regions,Terminator Regions,Genetic Terminator Region,Genetic Terminator Regions,Region, Genetic Terminator,Region, Terminator,Region, Transcriptional Terminator,Regions, Genetic Terminator,Regions, Terminator,Regions, Transcriptional Terminator,Sequence, Terminator,Sequences, Terminator,Terminator Region,Terminator Region, Genetic,Terminator Region, Transcriptional,Terminator Regions, Transcriptional,Terminator Sequences,Transcriptional Terminator Region