Biomaterial Database

Biochemical construction of specific chimeric plasmids from ColE1 DNA and unfractionated Escherichia coli DNA. 1976

C J Collins, and D A Jackson, and A J DeVries

A series of chimeric plasmids was constructed using colicinigenic factor E1 (ColE1) DNA as the replicon and DNA fragments carrying the galactose or tryptophan operons from E. coli. Restriction endonuclease EcoRI digests of ColE1 DNA and various DNAs containing the trp or gal operons were joined by T4 polynucleotide ligase [polynucleotide synthetase (ATP), poly(deoxyribonucleotide):poly(deoxyribonucleotide) ligase (AMP-forming), EC 6.5.1.1]. Chimeric plasmids carrying the desired genes were selected after transformation of Trp- or Gal- cells with ligated DNA. By using this method, we constructed ColE1-gal and ColE1-trp chimeric plasmids in which the source of the bacterial gal and trp operons was an unfractionated EcoRI digest of total E. coli DNA. The frequency of recovery of such chimeric plasmids is 10 to 20 colonies per mug of ligated DNA used in the transformation step. The method utilized in this report for constructing specific chimeric plasmids from total E. coli DNA is very simple. It requires only endonuclease R-EcoRI and T4 polynucleotide ligase, both of which are commercially available. The yield of transformants suggests that this method will be useful for cloning and amplifying a wide variety of functionally defined genes from E. coli and other prokaryotic organisms.

UI	MeSH Term	Description	Entries
D008970	Molecular Weight	The sum of the weight of all the atoms in a molecule.	Molecular Weights,Weight, Molecular,Weights, Molecular
D009876	Operon	In bacteria, a group of metabolically related genes, with a common promoter, whose transcription into a single polycistronic MESSENGER RNA is under the control of an OPERATOR REGION.	Operons
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
D011118	Polynucleotide Ligases	Catalyze the joining of preformed ribonucleotides or deoxyribonucleotides in phosphodiester linkage during genetic processes. EC 6.5.1.	Polynucleotide Synthetases,Ligases, Polynucleotide,Synthetases, Polynucleotide
D003087	Colicins	Bacteriocins elaborated by strains of Escherichia coli and related species. They are proteins or protein-lipopolysaccharide complexes lethal to other strains of the same species.	Colicin,Colicin E9,Colicine,Colicines,Colicin A,Colicin B,Colicin E,Colicin E1,Colicin E2,Colicin E3,Colicin E8,Colicin HSC10,Colicin Ia,Colicin Ib,Colicin K,Colicin K-K235,Colicin M,Colicin N,Colicin V,Colicins E,Colicins E9,Precolicin E1,Colicin K K235,E9, Colicin
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
D004269	DNA, Bacterial	Deoxyribonucleic acid that makes up the genetic material of bacteria.	Bacterial DNA
D004274	DNA, Recombinant	Biologically active DNA which has been formed by the in vitro joining of segments of DNA from different sources. It includes the recombination joint or edge of a heteroduplex region where two recombining DNA molecules are connected.	Genes, Spliced,Recombinant DNA,Spliced Gene,Recombinant DNA Research,Recombination Joint,DNA Research, Recombinant,Gene, Spliced,Joint, Recombination,Research, Recombinant DNA,Spliced Genes
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D005111	Extrachromosomal Inheritance	Vertical transmission of hereditary characters by DNA from cytoplasmic organelles such as MITOCHONDRIA; CHLOROPLASTS; and PLASTIDS, or from PLASMIDS or viral episomal DNA.	Cytoplasmic Inheritance,Extranuclear Inheritance,Inheritance, Cytoplasmic,Inheritance, Extrachromosomal,Inheritance, Extranuclear