Fungal protoplast fusion - a revisit
ABSTRACT Protoplast fusion is a non-specific recombination technique used for transfer of cytosolic organelles including genetic material. The process involves cell wall breakdown, regeneration of protoplasts, chemofusion and electrofusion. This review article discusses all the stages involved in fusion of protoplasts and some of the applications of protoplast fusion technique in fungal systems.
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ABSTRACT: Inactivated protoplast fusion by UV irradiation and UV+LiCl mutation was conducted using Nodulisporium sylviforme strain UV 40-19 and UL 50-6 to breed a high taxol-producing fungus. Qualitative and quantitative analysis of taxol production was confirmed using thin-layer chromatography, high performance liquid chromatography and mass spectrometry. The protoplasts of UV 40-19 and UL 50-6 were fully inactivated by heating at 54°C for 5 min and by UV irradiation (30 w UV light and vertical distance 30 cm) for 85 s. The highest fusion rate (14.31 ± 1.13%) between UV 40-19 and UL 50-6 was obtained under the conditions of 35% PEG, 90 s fusion time and the addition of 0.01 mol/l CaCl 2 . One high taxol production strain HDF-68 was obtained. The taxol production was up to 468.62 ± 37.49 g/l, which was increased by 24.51 and 19.35% compared with the parental strain UV 40-19 and UL 50-6 , respectively. This study provided a good basis for the application of this technique to the breeding of the strains with high taxol output.
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ABSTRACT: Paclitaxel is a potent and widely used antitumor agent. Considerable worldwide research efforts have been carried out on different production alternatives. Since the description of the first paclitaxel-producing fungi, more than 15 years ago, microorganisms have been investigated as potential alternatives for an environmentally acceptable, relatively simple and inexpensive method to produce paclitaxel. However, in spite of significant research on paclitaxel-producing microorganisms, no commercial fermentation process has been implemented up to now. The aim of this study is to review the present status of research on paclitaxel-producing microorganisms and the ongoing efforts to develop heterologous paclitaxel biosynthesis, and analyze the perspectives of microbial fermentation for paclitaxel production.Keywords: bacteria; fungi; genes; HPLC-MS; taxolThe Journal of Antibiotics 07/2010; 63(8):460-467. DOI:10.1038/ja.2010.83 · 2.04 Impact Factor
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ABSTRACT: An attempt was made to improve the stability toward centrifugation of protoplast fusion between Shewanella sp. and Escherichia coli. Stability of the cell membrane is an important factor in protoplast fusion. In order to change the fatty acid composition of the cell membrane phospholipids, eight fatty acids [caprylic acid, capric acid, palmitic acid, oleic acid, linoleic acid, linolenic acid, eicosapentaenoic acid (EPA) and docosahexaenoic acid] were added to each nutrient medium of Shewanella sp. and E. coli. The protoplasts were treated with lysozyme, and fusion occurred in the presence of a polyethylene glycol solution. The stability of the protoplast of Shewanella sp. decreased after EPA was added to the culture medium, and the stability of the protoplast of E. coli increased after the addition of linoleic acid or linolenic acid. Some fusant colonies that developed on the regenerated medium selected for E. coli with antibiotic tolerance. The efficiency of this fusion was higher than that of initial condition using protoplasts from Shewanella sp. and E. coli incubated without fatty acids. Protoplasts improved the fatty acid composition of the phospholipids. Cell membrane stability can change in order for the weak cells to be taken in by strong cells. These results suggested that the fatty acid composition of cell membrane phospholipids affected the fusant yield of the fusion of these bacteria.Fisheries Science 12/2008; 74(6):1290 - 1296. DOI:10.1111/j.1444-2906.2008.01654.x · 0.86 Impact Factor