PHYSIOLOGICAL AND GENETIC FACTORS AFFECTING TRANSFORMATION OF
            <i>BACILLUS SUBTILIS</i>

Young, Frank E.; Spizizen, John

doi:10.1128/jb.81.5.823-829.1961

Cited by 147 publications

(64 citation statements)

References 7 publications

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“…Plasmid DNA was routinely extracted using Plasmid MiniPrep Kit (Qiagen), whilst the extraction of B. subtilis chromosomal DNA was either done using DNA wizard kit (Promega) for sequencing and PCR, or as a simple DNA extract (47) to permit strain construction. The transformation of B. subtilis strains was carried out according to the method developed by Anagnostopoulos and Spizizen (48), modified by Young and Spizizen (49) and E. coli transformation was done as described by Hanahan,1985 (50). Selection for antibiotic resistance markers was done using the following concentrations: chloramphenicol (5 µg/ml), erythromycin (1 µg/ml), spectinomycin and ampicillin (100 µg/ml) in Nutrient Agar (Oxoid).…”

Section: General Materials and Methodsmentioning

confidence: 99%

Alanine Metabolism inBacillus subtilis

Sidiq

Chow

Zhao

et al. 2019

Preprint

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Despite a long history of genetic manipulation of Bacillus subtilis using auxotrophic markers in genetic manipulation, the genes involved in alanine metabolism have not been characterised fully. Here we show that B. subtilis expresses an alanine uptake permease, YtnA (DatA), that has a major role in the assimilation of D-alanine from the environment. Since this isomer of the amino acid is not normally abundant it likely source is form the cells own cell wall probably through the action of carboxypeptidases and/or the spontaneous release of D-alanine from the teichoic acids. Also in this work we clarify the synthetic pathways acting in the biosynthesis of alanine. Genetically we show that, unlike E. coli where multiple enzymes have a biochemical activity that can generate alanine, in B. subtilis the primary synthetic enzyme for alanine is encoded by alaT, although a second gene, dat, is present that can support slow growth of an alanine auxotroph however our data suggests that this enzyme probably synthesises D-alanine. In summary this work has provided an explanation of the observation that growth of B. subtilis is linked with an efficient recycling system for D-alanine that is released from the cell as the cell envelope is processed to permit cell enlargement. The results also suggest that the relative abundance of D- and L-alanine that might be linked with cytosolic pool of D and L-glutamate, and so enabling tight coupling protein and cell envelope synthesis with the metabolic status of the cell.

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Section: General Materials and Methodsmentioning

confidence: 99%

Alanine Metabolism inBacillus subtilis

Sidiq

Chow

Zhao

et al. 2019

Preprint

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“…Broth Transformation. -Strains of B. subtilis were grown to competence for transformation by the two-step procedure of Young and Spizizen (1961). Cells of strain M28 were grown to competence using the procedures of Thorne and Stull (1966) and McCuen and Thorne (1971).…”

Section: Laboratory Strainsmentioning

confidence: 99%

Genetic Exchange between Bacillus subtilis and Bacillus licheniformis: Variable Hybrid Stability and the Nature of Bacterial Species

Duncan¹,

Istock²,

Graham³

et al. 1989

Evolution

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Experiments employing both broth and soil cultures demonstrated the capacity for bidirectional genetic exchange between the eubacterial species Bacillus subtilis and Bacillus licheniformis. The process was studied using standard laboratory strains and wild isolates of these species. The genetic exchange in soil occurs spontaneously. The interspecific recombination involved markers for antibiotic resistance and for the use of specific carbon sources (API characters). Hybrids frequently had unstable phenotypes, i.e., lacked a consistent expression of foreign genes over repeated transfer and growth. This instability often involved a "correction" back toward the phenotype of one or the other of the parental species for many differentiating characters; the final phenotype was always that of the more probable or actually known recipient species. This "correction" process is reminiscent of phenomena associated with the instability of artificial fusion protoplasts or noncomplementing diploids of B. subtilis, as well as the merodiploids formed by intergeneric crosses with enteric bacteria. The hybrids observed here must also be diploid, in some manner, because they sequentially express traits of both parental species at rates well above the frequency of mutation. Among the unstable changes in hybrids of the wild strains there was a 3:1 bias in favor of "correction." The dynamics of the hybridization process in soil are described. It appears that the hybrids are formed most rapidly following outgrowth from spores and during the early growth of parental vegetative cell populations. Later on, the hybrids are much less frequent in the soil cultures, suggesting that they are competitively inferior to the parental species. It is argued that the capacity for recombination found between B. subtilis and B. licheniformis could locally erase their distinctness, even though they possess only about 15% DNA sequence homology. Yet they remain distinct in the wild. The methods and results of these experiments prepare the way for detailed studies of the nature of species and species boundaries throughout the genus Bacillus.

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“…[Occasional tfmi strains are similarly found among various auxotrophic mutants; this may reflect the fact that loss of transformability may have more than one cause. The claim that spmutants cannot be transformed (206,238) must have been based on the examination of only a few strains.] (v) Several SPOAstrains are affected in the lyt character [not all of them, however, as was first thought (175)], and some in the ms trait.…”

Section: Cytological Classification Of the Mutantsmentioning

confidence: 99%