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[Study on Bacillus pumilus as a recipient strain for genetic engineering of Bacillus]
Abstract:
Bacillus pumilus 289 can be transformed easily by plasmid pUB110 through protoplast transformation with the frequency of 10(-5)--10(-3), similar to B. subtilis AS 1.1176, a derivative strain of B. subtilis 168. The regeneration frequency of its protoplast is only slightly lower than B. subtilis AS1.1176 (0.3-12.0% compare to 1.53--24.16%). Plasmid pUB110 can be maintained in both bacterial strains stably. The frequency of loss of the plasmid in both strains is lower than 3% after 45 generations in LB medium. But it is quite different that the hybrid plasmid (pUB110 with 3.9 kb foreign DNA fragment) can be maintained much more stably in B. pumilus 289 than in B. subtilis AS1.1176. The frequency of loss of the plasmid is lower than 5% in B. pumilus 289 and 24% in B. subtilis AS1.1176 after 25 generations when they grown in SH medium. The expression level of foreign gene in B. pumilus 289 is also much higher than that in B. subtilis AS1.1176. Therefore B. pumilus 289 is valuable to be exploited as recipient strain for genetic engineering of Bacillus in the future.
Insights
Bacillus pumilus 289 shows efficient plasmid transformation and stable maintenance, outperforming Bacillus subtilis in hybrid plasmid stability and foreign gene expression. This makes B. pumilus 289 a promising candidate for future genetic engineering applications.
Area of Science:
- Microbiology
- Molecular Biology
- Biotechnology
Background:
- Bacillus subtilis is a well-established host for genetic engineering.
- Exploring alternative Bacillus species for enhanced genetic manipulation is crucial.
- Bacillus pumilus 289 is a potential candidate for biotechnological applications.
Purpose of the Study:
- To evaluate Bacillus pumilus 289 as a host for genetic engineering.
- To compare its transformation efficiency and plasmid stability with Bacillus subtilis.
- To assess the expression of foreign genes in B. pumilus 289.
Main Methods:
- Protoplast transformation using plasmid pUB110.
- Assessing protoplast regeneration frequency.
- Evaluating plasmid stability and loss frequency over generations.
- Measuring foreign gene expression levels.
Main Results:
- Bacillus pumilus 289 demonstrated efficient transformation with plasmid pUB110 (10(-5)--10(-3) frequency).
- Plasmid pUB110 exhibited stable maintenance in B. pumilus 289 with low loss rates (<3% after 45 generations).
- Hybrid plasmids were significantly more stable in B. pumilus 289 than in Bacillus subtilis, with lower loss rates (<5% vs. 24% after 25 generations).
- Foreign gene expression was markedly higher in B. pumilus 289 compared to B. subtilis.
Conclusions:
- Bacillus pumilus 289 is an effective recipient strain for plasmid transformation.
- B. pumilus 289 offers superior stability for hybrid plasmids and enhanced foreign gene expression.
- B. pumilus 289 holds significant potential as a valuable host for future Bacillus genetic engineering endeavors.