Related Experiment Videos
Isolation and characterization of a unique division mutant of Bacillus megaterium
D A Lach1, V K Sharma, P S Vary
1Department of Biological Sciences, Northern Illinois University, DeKalb, 60115.
Abstract:
A filamentous division mutant, PV302, of Bacillus megaterium QM B1551 was isolated while screening for sporulation-defective mutants after nitrosoguanidine mutagenesis. Both phase-contrast and electron microscopy revealed that the mutant produced small spherical cells as well as filaments. It also accumulated large amounts of poly-beta-hydroxybutyrate. Poly-beta-hydroxybutyrate accounted for 16% of the dry weight of the mutant strain even after 28 h growth. In comparison to the parental strain, the division mutant also showed both an inability to sporulate and a reduced growth rate. All these phenotypes transduced together. Revertants gained the ability to sporulate, divide, and grow normally. Transductional mapping of the mutation, designated div-1, established a new linkage group for B. megaterium consisting of div-1 and the pyrimidine biosynthesis genes pyrD BCF. The spherical cells were separated from filaments by sucrose gradients and were tested for nucleic acid content and viability. The purified spherical cell fraction contained one-fifth the amount of DNA per mg protein as compared with the filamentous cell fraction and was shown to contain both non-viable minicells and some cells capable of growing after a lag of about 4 h. This suggests that the mutation not only causes defects in septum placement and sporulation, but may possibly affect DNA partitioning.
Insights
A Bacillus megaterium mutant exhibits defects in cell division, sporulation, and DNA partitioning, accumulating poly-beta-hydroxybutyrate and producing minicells. This div-1 mutation establishes a new linkage group in this bacterium.
Area of Science:
- Microbiology
- Bacterial Genetics
- Cell Biology
Background:
- Screening for sporulation-defective mutants in Bacillus megaterium QM B1551 led to the isolation of a filamentous division mutant, PV302.
- Nitrosoguanidine mutagenesis was employed to generate mutations affecting bacterial growth and development.
Purpose of the Study:
- To characterize a novel Bacillus megaterium mutant (PV302) exhibiting defects in cell division and sporulation.
- To investigate the genetic basis and phenotypic consequences of the div-1 mutation.
Main Methods:
- Phase-contrast and electron microscopy were used to visualize cellular morphology.
- Biochemical analysis quantified poly-beta-hydroxybutyrate accumulation.
- Transductional mapping and sucrose gradient separation were employed for genetic and cellular analysis.
Main Results:
- The mutant PV302 produced filaments and small spherical cells, accumulating significant poly-beta-hydroxybutyrate (16% of dry weight).
- PV302 demonstrated an inability to sporulate, a reduced growth rate, and produced minicells with reduced DNA content.
- Transductional mapping placed the div-1 mutation in a new linkage group with pyrimidine biosynthesis genes.
Conclusions:
- The div-1 mutation in Bacillus megaterium affects septum placement, sporulation, and potentially DNA partitioning, leading to minicell formation.
- The study establishes a new linkage group for B. megaterium, linking div-1 to pyrD BCF.
- The mutant's phenotypes, including poly-beta-hydroxybutyrate accumulation and impaired division, highlight complex regulatory networks in bacterial development.