Incorporation of the Whole Chromosomal DNA in Protoplast Lysates into Competent Cells of Bacillus subtilis

  title={Incorporation of the Whole Chromosomal DNA in Protoplast Lysates into Competent Cells of Bacillus subtilis},
  author={Takashi Akamatsu and Hisataka Taguchi},
  journal={Bioscience, Biotechnology, and Biochemistry},
  pages={823 - 829}
Competent cells of Bacillus subtilis AC870 (purB, leuB, trpC, ald-1) were transformed to Ade+, Trp+, or Ade+ Trp+ with DNA in protoplast lysates of B. subtilis AC819 (hisH, tet-1, rpsL, smo-1). The contransfer ratio of purB to trpC was constant at 7-9% (Ade+ Trp+/Trp+) or 3% (Ade+ Trp+/Ade+) at protoplast concentrations of 2.7×103∼2.7×106 per ml. The whole chromosomal DNA must be certainly incorporated into competent cells from the following reasons; (1) purB is opposite to trpC on the… 

Fate of transforming bacterial genome following incorporation into competent cells of Bacillus subtilis: a continuous length of incorporated DNA.

The results strongly suggest that the length of continuous DNA incorporated into B. subtilis is most probably greater than 1271 kb, which is a working hypothesis on the mechanism of the transformation of competent B.subilis by DNA in protoplast lysates (LP transformation).

DNA taken into Bacillus subtilis competent cells by lysed-protoplast transformation is not ssDNA but dsDNA.

All of the observations strongly suggest that the incorporated DNA is dsDNA, and the transformation of competent B. subtilis by DNA in protoplast lysate is different from that by purified DNA taken up conventionally.

Functional linkages between replication proteins of genes 1, 3 and 5 of Bacillus subtilis phage φ29.

It is confirmed that the absence of gp1 at non-permissive temperature can be compensated by the suppressors which have the single amino acid substitution in either gp5 or gp3, and proposed its new regulatory subdomain at which other molecules including gp1 would interact with and regulate functions of gp3.

The accurate replacement of long genome region more than several hundreds kilobases in Bacillus subtilis.

Competent cell transformation with DNA obtained by the gentle lysis of protoplasts (LP transformation) was used to replace a large genomic region in this study. Discontinuity was detected in the

Optimization of a whole-cell biocatalyst by employing genetically encoded product sensors inside nanolitre reactors.

The underlying design principles of the assay are general and enable the development of similar protocols, which ultimately will speed up the optimization of whole-cell biocatalysts.

How Many Genes Does a Cell Need

This chapter reviews, discusses, and proposes studies aimed at determining how many Mycoplasma genitalium genes are really necessary when cells are grown under ideal laboratory conditions, including recA, which is one of the most ubiquitous proteins found in nature.



Interspecific Transformation of Bacillus subtilis Competent Cells by Chromosomal DNA in Lysates of Protoplasts of Bacillus amyloliquefaciens

The involvement of mutS in the interspecific transformation was not significant and an in- terspecific cotransformation ratio as high as an intraspecific one using purified DNA was also detected between cysA and rpsL markers, which are separated by 16 kb on the B. subtilis chromosome.

Number of Deoxyribonucleic Acid Uptake Sites in Competent Cells of Bacillus subtilis

  • R. Singh
  • Biology, Chemistry
    Journal of bacteriology
  • 1972
Two direct methods are presented for estimating the average number of deoxyribonucleic acid (DNA) uptake sites in competent cells of Bacillus subtilis from measurement of 14C- or 3H-thymine-labeled

Association of the Bacillus subtilis Chromosome with the Cell Membrane: Resolution of Free and Bound Deoxyribonucleic Acid on Renografin Gradients

Linear density gradients of Renografin have resolved two components of bacterial deoxyribonucleic acid in sheared lysates and suggest that replicating DNA at the origin and possibly the terminus of replication are associated with membrane.

Molecular cloning and sequence of comK, a gene required for genetic competence in Bacillus subtilis

Epistatic interactions indicate that comK is a competence locus occupying an intermediate position in the competence signal transduction network in Bacillus subtilis.

Characterization of comE, a late competence operon of Bacillus subtilis required for the binding and uptake of transforming DNA

The characterization of comE is presented, an operon under competence control that is required for both DNA binding to the competent ceil surface, and for uptake of DNA by competent Bacillus subtilis.

Reexamination of phenotypic defects in rec-1 and rec-2 mutants of Haemophilus influenzae Rd

Radiolabeled donor DNA is efficiently taken up into competent H. influenzae Rd rec-2 mutant cells but does not undergo the rapid degradation observed in wild-type cells, and donor label is not recovered in the chromosome even after 1 h, interpreted as a failure of the donor DNA to be translocated out of the transformasome.

An Improved Method of Protoplast Regeneration for Bacillus Species and Its Application to Protoplast Fusion and Transformation

An improved method for regenerating Bacillus subtilis protoplasts at the frequency of 92∼100% on a semi-synthetic medium was found. Protoplasts were preincubated in HCP-3 medium, an isotonic

comK acts as an autoregulatory control switch in the signal transduction route to competence in Bacillus subtilis

It is shown that expression of comK is dependent on its own gene product as well as on the gene products of all other tested regulatory genes known to be involved in competence development and infer that the signals which trigger competence development, after having been received and processed by the various components of the competence signal transduction pathway, all converge at the level ofcomK expression.

The regulation of competence transcription factor synthesis constitutes a critical control point in the regulation of competence in Bacillus subtilis

Various competence regulatory genes (comA, srfA, degU, abrB, sin, and spo0A) are shown to be required for the expression of comK and srfA transcription is shown to occur equally in cells destined for competence and those destined not to become competent.