An efficient transformation method for Bacillus subtilis DB104.
Summary of "An efficient transformation method for Bacillus subtilis DB104."
Bacillus subtilis strains are used for extracellular expression of enzymes (i.e., proteases, lipases, and cellulases) which are often engineered by directed evolution for industrial applications. B. subtilis DB104 represents an attractive directed evolution host since it has a low proteolytic activity and efficient secretion. B. subtilis DB104 is hampered like many other Bacillus strains by insufficient transformation efficiencies (≤10(3) transformants/μg DNA). After investigating five physical and chemical transformation protocols, a novel natural competent transformation protocol was established for B. subtilis DB104 by optimizing growth conditions and histidine concentration during competence development, implementing an additional incubation step in the competence development phase and a recovery step during the transformation procedure. In addition, the influence of the amount and size of the transformed plasmid DNA on transformation efficiency was investigated. The natural competence protocol is "easy" in handling and allows for the first time to generate large libraries (1.5 × 10(5) transformants/μg plasmid DNA) in B. subtilis DB104 without requiring microgram amounts of DNA.
Affiliation
Lehrstuhl für Biotechnologie, RWTH Aachen University, Worringerweg 1, 52074, Aachen, Germany.
Journal Details
This article was published in the following journal.
Name: Applied microbiology and biotechnology
ISSN: 1432-0614
Pages:
Links
- PubMed Source: http://www.ncbi.nlm.nih.gov/pubmed/22395911
- DOI: http://dx.doi.org/10.1007/s00253-012-3987-2
Medical and Biotech [MESH] Definitions
Subtilisins
A family of SERINE ENDOPEPTIDASES isolated from Bacillus subtilis. EC 3.4.21.-
Bacillus Subtilis
A species of gram-positive bacteria that is a common soil and water saprophyte.
Mycobacillin
A cyclic polypeptide antibiotic isolated from culture filtrates of Bacillus subtilis that acts as an antifungal agent.
Subtilisin
A serine endopeptidase isolated from Bacillus subtilis. It hydrolyzes proteins with broad specificity for peptide bonds, and a preference for a large uncharged residue in P1. It also hydrolyzes peptide amides. (From Enzyme Nomenclature, 1992) EC 3.4.21.62.
Bacitracin
A complex of cyclic peptide antibiotics produced by the Tracy-I strain of Bacillus subtilis. The commercial preparation is a mixture of at least nine bacitracins with bacitracin A as the major constituent. It is used topically to treat open infections such as infected eczema and infected dermal ulcers. (From Goodman and Gilman, The Pharmacological Basis of Therapeutics, 8th ed, p1140)
PubMed Articles
A modified electroporation method using trehalose is presented for the transformation of Bacillus subtilis. The new method improved the transformation efficiency of B. subtilis nearly 2,000-fold compa...
Chemostat-induced uneven division of Bacillus subtilis.
Anomalous forms of Bacillus subtilis A32 produced by prolonged cultivation in a chemostat under nitrogen limitation are described. A change in the cultivation conditions brought about a transformation...
Bacillus subtilis SC-8 is a Gram-positive bacterium displaying narrow antagonistic activity for the Bacillus cereus group. B. subtilis SC-8 was isolated from Korean traditional fermented-soybean food....
Genome of a Gut Strain of Bacillus subtilis.
Bacillus subtilis is a Gram-positive, rod-shaped, spore-forming bacterium. We present the genome sequence of an undomesticated strain, BSP1, isolated from poultry. The sequence of the BSP1 genome supp...
The gene encoding a novel alkaline pectate lyase (Apel) from Bacillus subtilis was cloned and expressed in B. subtilis WB600. Apel contained an ORF of 1,260 bp, encoding a signal peptide of 21 amino...
Clinical Trials
DIAMANTE - Diseases Infectious Analysis Microflora Antibiotic Effect
To assess the effects of Bacillus clausii (Enterogermina®) on fecal microbial flora (using Polymerase Chain Reaction - Denaturing Gradient Gel Electrophoresis - PCR-DGGE - Method) in ant...
TEDDY: Spores of Bacillus Clausii in Acute Diarrhoea in Children
Primary objective: - To demonstrate the efficacy of a Bacillus clausii probiotic strain compared to placebo in children suffering from acute diarrhoea and treated for 7 days....
Preventing Antibiotic-Associated DiarRhea Using Erceflora
To determine the effectiveness of the pre-biotic Bacillus clausii in preventing antibiotic associated diarrhea among hospitalized immunocompetent Filipino children.
The goal of this clinical research study is to find the highest tolerable dose of fludarabine and cytarabine that can be given in combination with oxaliplatin and rituximab in the treatmen...
Diathermy Cone Biopsy: A Study of Two Techniques.
The purpose of this trial is to determine whether SWETZ (Straight Wire Excision of Transformation Zone) is a superior alternative to LLETZ cone biopsy in the management of pre-invasive end...
