Methods

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Here you can find a list of methods for working with Bacillus subtilis:

Media

A collection of mutant strains

  • the complete mutant strain collection from the Koo et al. paper is now available: get access here

Toolboxes for the work with B. subtilis

  • the biobrick box PubMed
  • a part toolbox to tune genetic expression PubMed

Analysis of protein-protein interactions

  • Bacterial Two Hybrid: BACTH

Analysis of membrane proteins

  • application of a split green fluorescent protein reporter PubMed

Antibodies

Protein expression

Strain construction

  • introduction of markerfree deletions: PubMed
  • generation of markerfree mutations: PubMed
  • an improved method for transformation: PubMed

RNA analysis

  • Mechanical cell disruption and extraction of RNA from B. subtilis: Media:SOP-RNA.pdf

Genetic work with non-standard strains

  • transformation of wild-type B. subtilis strains incl. NCIB3610: PubMed
  • efficient electroporation of B. subtilis PubMed
  • a novel transformation protocol for B. subtilis DB104 PubMed

General procedures

  • Standard operation procedures (SOPs) of the SYSMO-BACELL consortium

Microscopic techniques

  • Live cell imaging of B. subtilis cells using automated time-lapse microscopy PubMed
  • Assay of gene expression dynamics using live cell imaging PubMed
  • Visualization and quantification of gene expression heterogeneity in growing microbial cells PubMed
  • GFP variants specifically optimizd for use in B. subtilis PubMed
  • Quantitative analysis of cell types by comparing microscoy images PubMed
  • Semi-automated single cell analysis using membrane or DNA staining PubMed

Transposon mutagenesis

  • mariner transposon mutagenesis for random inducible-expression insertions and transcriptional reporter fusion insertions PubMed

Protein-DNA interactions

  • ChAP-chip: A modified ChIP-chip protocol for the in vivo identification of binding sites of DNA-binding proteins PubMed

Genome engineering

  • generalized bacterial genome editing using mobile group II introns and Cre-lox PubMed
  • genome engineering using a synthetic gene circuit PubMed
  • Ordered Gene Assembly in Bacillus subtilis (OGAB) PubMed
  • construction of a super-competent strain PubMed
  • use of the CRISPR-Cas system PubMed
  • A review on genome engineering:


Visual descriptions of methods for B. subtilis in the Journal of Visual Experimentation

Lorena Stannek, Richard Egelkamp, Katrin Gunka, Fabian M Commichau
Monitoring intraspecies competition in a bacterial cell population by cocultivation of fluorescently labelled strains.
J Vis Exp: 2014, (83);e51196
[PubMed:24473333] [WorldCat.org] [DOI] (I e)

Samantha M Desmarais, Felipe Cava, Miguel A de Pedro, Kerwyn Casey Huang
Isolation and preparation of bacterial cell walls for compositional analysis by ultra performance liquid chromatography.
J Vis Exp: 2014, (83);e51183
[PubMed:24457605] [WorldCat.org] [DOI] (I e)

Ewa Król, Dirk-Jan Scheffers
FtsZ polymerization assays: simple protocols and considerations.
J Vis Exp: 2013, (81);e50844
[PubMed:24300445] [WorldCat.org] [DOI] (I e)

Elizabeth Anne Shank
Using coculture to detect chemically mediated interspecies interactions.
J Vis Exp: 2013, (80);e50863
[PubMed:24300024] [WorldCat.org] [DOI] (I e)

Juan C Garcia-Betancur, Ana Yepes, Johannes Schneider, Daniel Lopez
Single-cell analysis of Bacillus subtilis biofilms using fluorescence microscopy and flow cytometry.
J Vis Exp: 2012, (60);
[PubMed:22371091] [WorldCat.org] [DOI] (I e)

Imke G de Jong, Katrin Beilharz, Oscar P Kuipers, Jan-Willem Veening
Live Cell Imaging of Bacillus subtilis and Streptococcus pneumoniae using Automated Time-lapse Microscopy.
J Vis Exp: 2011, (53);
[PubMed:21841760] [WorldCat.org] [DOI] (I e)

Andrew D Klocko, Kaleena M Crafton, Brian W Walsh, Justin S Lenhart, Lyle A Simmons
Imaging mismatch repair and cellular responses to DNA damage in Bacillus subtilis.
J Vis Exp: 2010, (36);
[PubMed:20142799] [WorldCat.org] [DOI] (I e)


Key references