Biomaterial Database

T7 RNA polymerase-driven inducible cell lysis for DNA transfer from Escherichia coli to Bacillus subtilis. 2017

Mario Juhas, and James W Ajioka

Department of Pathology, University of Cambridge, Tennis Court Road, CB2 1QP, Cambridge, UK.

The majority of the good DNA editing techniques have been developed in Escherichia coli; however, Bacillus subtilis is better host for a plethora of synthetic biology and biotechnology applications. Reliable and efficient systems for the transfer of synthetic DNA between E. coli and B. subtilis are therefore of the highest importance. Using synthetic biology approaches, such as streamlined lambda Red recombineering and Gibson Isothermal Assembly, we integrated genetic circuits pT7L123, Repr-ts-1 and pLT7pol encoding the lysis genes of bacteriophages MS2, ΦX174 and lambda, the thermosensitive repressor and the T7 RNA polymerase into the E. coli chromosome. In this system, T7 RNA polymerase regulated by the thermosensitive repressor drives the expression of the phage lysis genes. We showed that T7 RNA polymerase significantly increases efficiency of cell lysis and transfer of the plasmid and bacterial artificial chromosome-encoded DNA from the lysed E. coli into B. subtilis. The T7 RNA polymerase-driven inducible cell lysis system is suitable for the efficient cell lysis and transfer of the DNA engineered in E. coli to other naturally competent hosts, such as B. subtilis.

UI	MeSH Term	Description	Entries
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
D004269	DNA, Bacterial	Deoxyribonucleic acid that makes up the genetic material of bacteria.	Bacterial DNA
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
D001412	Bacillus subtilis	A species of gram-positive bacteria that is a common soil and water saprophyte.	Natto Bacteria,Bacillus subtilis (natto),Bacillus subtilis subsp. natto,Bacillus subtilis var. natto
D001435	Bacteriophages	Viruses whose hosts are bacterial cells.	Phages,Bacteriophage,Phage
D012321	DNA-Directed RNA Polymerases	Enzymes that catalyze DNA template-directed extension of the 3'-end of an RNA strand one nucleotide at a time. They can initiate a chain de novo. In eukaryotes, three forms of the enzyme have been distinguished on the basis of sensitivity to alpha-amanitin, and the type of RNA synthesized. (From Enzyme Nomenclature, 1992).	DNA-Dependent RNA Polymerases,RNA Polymerases,Transcriptases,DNA-Directed RNA Polymerase,RNA Polymerase,Transcriptase,DNA Dependent RNA Polymerases,DNA Directed RNA Polymerase,DNA Directed RNA Polymerases,Polymerase, DNA-Directed RNA,Polymerase, RNA,Polymerases, DNA-Dependent RNA,Polymerases, DNA-Directed RNA,Polymerases, RNA,RNA Polymerase, DNA-Directed,RNA Polymerases, DNA-Dependent,RNA Polymerases, DNA-Directed
D014764	Viral Proteins	Proteins found in any species of virus.	Gene Products, Viral,Viral Gene Products,Viral Gene Proteins,Viral Protein,Protein, Viral,Proteins, Viral
D058615	Synthetic Biology	A field of biological research combining engineering in the formulation, design, and building (synthesis) of novel biological structures, functions, and systems.	Biologies, Synthetic,Biology, Synthetic,Synthetic Biologies
D061269	DNA Transformation Competence	The ability of bacterial cells to take up exogenous DNA and be genetically transformed by it.	Competence, DNA Transformation,Transformation Competence, DNA