Protoplast Transformation of Recalcitrant Alkaliphilic Bacillus sp. with Methylated Plasmid DNA and a Developed Hard Agar Regeneration Medium

Gao, Chenghua; Xue, Yanfen; Ma, Yanhe

doi:10.1371/journal.pone.0028148

Cited by 16 publications

(19 citation statements)

References 49 publications

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“…Most Bacillus species, regardless of their source, harbour at least one indigenous plasmid (Molina-Aja et al, 2002;Reyaz et al, 2013;Lobova et al, 2015). These bacterial isolates have further corroborated the submission of Gao et al (2011) that Bacillus spp., have evolved several genetic mediated mechanisms to survive in extreme environment.…”

Section: Resultssupporting

confidence: 61%

Plasmid Profiles of Extremophiles Associated with Indigenous Black Soaps and Functional Groups Present

Oyeniran¹,

Oladunmoye²

2015

Microbiology J.

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In the present study, plain indigenous black soap market sample and black soap fortified with brown and white eggshell powder effectively at 20 g/250 mL of molten soap were analyzed microbiologically using standard method after which the bacterial isolates were profiled for presence of plasmid. Structural elucidation of the black soaps was achieved by Gas Chromatography Mass Spectrometry (GCMS) analysis of the methylated fraction. Fungal isolates from plain black soap were Chrysosporium spp. and Aspergillus granulosus while the bacterial isolate: Bacillus vedderi contained plasmid with an estimated molecular weight of 17578 bp. Ramulispora javanicus and Aspergilus flavus while Bacillus vedderi and Bacillus faraginis were fungal isolates were also recorded from black soap fortified with brown and white eggshell powder, respectively. The GCMS analysis revealed fatty acid as the predominant functional group. Plasmid presence in these bacteria may confer extremophily status on them and thus aid their survival in this environment. Antimicrobial activities of the indigenous black in folkloric applications may be predicated on the presence of its fatty acid compounds.

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Section: Resultssupporting

confidence: 61%

Plasmid Profiles of Extremophiles Associated with Indigenous Black Soaps and Functional Groups Present

Oyeniran¹,

Oladunmoye²

2015

Microbiology J.

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“…Although several thermophilic Bacillus species have been described to be genetically accessible (11,24,29,31,(37)(38)(39), often laborious protocols, such as protoplast transformation, are required. Moreover, for each species, highly variable transformation efficiencies have been reported for a limited number of strains, suggesting that genetic accessibility is very strain specific.…”

Section: Discussionmentioning

confidence: 99%

Isolation and Screening of Thermophilic Bacilli from Compost for Electrotransformation and Fermentation: Characterization of Bacillus smithii ET 138 as a New Biocatalyst

Bosma

Weijer

Daas

et al. 2015

Appl Environ Microbiol

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b Thermophilic bacteria are regarded as attractive production organisms for cost-efficient conversion of renewable resources to green chemicals, but their genetic accessibility is a major bottleneck in developing them into versatile platform organisms. In this study, we aimed to isolate thermophilic, facultatively anaerobic bacilli that are genetically accessible and have potential as platform organisms. From compost, we isolated 267 strains that produced acids from C 5 and C 6 sugars at temperatures of 55°C or 65°C. Subsequently, 44 strains that showed the highest production of acids were screened for genetic accessibility by electroporation. Two Geobacillus thermodenitrificans isolates and one Bacillus smithii isolate were found to be transformable with plasmid pNW33n. Of these, B. smithii ET 138 was the best-performing strain in laboratory-scale fermentations and was capable of producing organic acids from glucose as well as from xylose. It is an acidotolerant strain able to produce organic acids until a lower limit of approximately pH 4.5. As genetic accessibility of B. smithii had not been described previously, six other B. smithii strains from the DSMZ culture collection were tested for electroporation efficiencies, and we found the type strain DSM 4216 T and strain DSM 460 to be transformable. The transformation protocol for B. smithii isolate ET 138 was optimized to obtain approximately 5 ؋ 10 3 colonies per g plasmid pNW33n. Genetic accessibility combined with robust acid production capacities on C 5 and C 6 sugars at a relatively broad pH range make B. smithii ET 138 an attractive biocatalyst for the production of lactic acid and potentially other green chemicals. Green chemicals are sustainable bio-based alternatives for chemicals based on fossil resources. Biomass is considered an attractive renewable resource for the production of such green chemicals. Major challenges for using biomass are to develop costeffective production processes and to convert substrates that go beyond first-generation pure sugars. From this perspective, the use of microbial fermentation processes that convert lignocellulosic sugars is gaining considerable attention. For particular products natural producers can be used, but often Escherichia coli and Saccharomyces cerevisiae are used as platform organisms because of their well-known physiology and ease of engineering for the production of many different products (1, 2).In general, most organisms used for the production of green chemicals and fuels are mesophiles, and current processes still are based mainly on first-generation feedstocks, i.e., sucrose from sugar beet or sugarcane or glucose derived from starches from corn or tapioca. Although the engineering of platform organisms broadens the spectrum of possible substrates (3), the efficiency of the process could be increased by the use of organisms naturally capable of degrading lignocellulose-derived sugars (4). To further increase the efficiency and reduce the costs of the microbial production of green chemicals, modera...

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“…It can utilize various types of sugars, such as pentoses, hexoses, and polysaccharides [ 17 , 18 ]. Importantly, a genetic manipulation system has been successfully developed for this promising strain [ 19 ]. Thus, these characteristics make Bacillus sp.…”

Section: Introductionmentioning

confidence: 99%

Efficient fermentative production of polymer-grade d-lactate by an engineered alkaliphilic Bacillus sp. strain under non-sterile conditions

Assavasirijinda

et al. 2016

Microb Cell Fact

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BackgroundPolylactic acid (PLA) is one important chemical building block that is well known as a biodegradable and a biocompatible plastic. The traditional lactate fermentation processes need CaCO3 as neutralizer to maintain the desired pH, which results in an amount of insoluble CaSO4 waste during the purification process. To overcome such environmental issue, alkaliphilic organisms have the great potential to be used as an organic acid producer under NaOH-neutralizing agent based fermentation. Additionally, high optical purity property in d-lactic acid is now attracting more attention from both scientific and industrial communities because it can improve mechanical properties of PLA by blending l- or d-polymer together. However, the use of low-price nitrogen source for d-lactate fermentation by alkaliphilic organisms combined with NaOH-neutralizing agent based process has not been studied. Therefore, our goal was the demonstrations of newly simplify high-optical-purity d-lactate production by using low-priced peanut meal combined with non-sterile NaOH-neutralizing agent based fermentation.ResultsIn this study, we developed a process for high-optical-purity d-lactate production using an engineered alkaliphilic Bacillus strain. First, the native l-lactate dehydrogenase gene (ldh) was knocked out, and the d-lactate dehydrogenase gene from Lactobacillus delbrueckii was introduced to construct a d-lactate producer. The key gene responsible for exopolysaccharide biosynthesis (epsD) was subsequently disrupted to increase the yield and simplify the downstream process. Finally, a fed-batch fermentation under non-sterile conditions was conducted using low-priced peanut meal as a nitrogen source and NaOH as a green neutralizer. The d-lactate titer reached 143.99 g/l, with a yield of 96.09 %, an overall productivity of 1.674 g/l/h including with the highest productivity at 16 h of 3.04 g/l/h, which was even higher than that of a sterile fermentation. Moreover, high optical purities (approximately 99.85 %) of d-lactate were obtained under both conditions.ConclusionsGiven the use of a cheap nitrogen source and a non-sterile green fermentation process, this study provides a more valuable and favorable fermentation process for future polymer-grade d-lactate production.

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Protoplast Transformation of Recalcitrant Alkaliphilic Bacillus sp. with Methylated Plasmid DNA and a Developed Hard Agar Regeneration Medium

Cited by 16 publications

References 49 publications

Plasmid Profiles of Extremophiles Associated with Indigenous Black Soaps and Functional Groups Present

Plasmid Profiles of Extremophiles Associated with Indigenous Black Soaps and Functional Groups Present

Isolation and Screening of Thermophilic Bacilli from Compost for Electrotransformation and Fermentation: Characterization of Bacillus smithii ET 138 as a New Biocatalyst

Efficient fermentative production of polymer-grade d-lactate by an engineered alkaliphilic Bacillus sp. strain under non-sterile conditions

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