Sub-cellular localization and overexpressing analysis of hydroxylase gene TcCYP725A22 of Taxus chinensis / 生物工程学报

The discovery of hydroxylases in the anticancer drug taxol biosynthesis pathway is a hotspot and difficulty in current research. In this study, a new hydroxylase gene TcCYP725A22 (GenBank accession number: MF448646.1) was used to construct a sub-cellular localization vector pCAMIBA1303-TcCYP725A22-EGFP to get the transient expression in onion epidermal cells. Laser confocal microscopy revealed that the protein encoded by this gene was localized in the cell membrane. Furthermore, the recombinant plant expression plasmid pBI121-TcCYP725A22 was constructed. After transient transformation to the Taxus chinensis mediated by Agrobacterium tumefaciens LBA4404, qRT-PCR and LC-MS were utilized to analyze the effects of TcCYP725A22 overexpression on the synthesis of taxol. The results showed that, in the TcCYP725A22 overexpressed cell line, expression levels of most defined hydroxylase genes for taxol biosynthesis were increased, and the yield of taxanes were also increased. It was concluded that the hydroxylase gene TcCYP725A22 is likely involved in the biosynthetic pathway of taxol.

Research progress in hydroxylase in taxol biosynthetic pathway / 生物工程学报

Taxol is a secondary metabolite with prominent anti-tumor activity, but the yield cannot meet the growing clinical demand due to lower content in yew. Now, most enzyme genes involved in taxol biosynthesis have been cloned and identified, so that obtaining this drug by using synthetic biology method has become a hotspot in recent years. However, most hydroxylases involved in taxol biosynthetic pathway have not been explored. Here, we reviewed the progress on the biosynthesis pathway of taxol, especially concerning hydroxylase. The future research areas of taxol biosynthesis through synthetic biology were also discussed to provide basis for the discovery of uncharacterized hydroxylase genes and the mass taxol production by synthetic biology technology.

Controllable bio-fabrication based on microbes / 生物工程学报

Microorganisms in nature have rich variety, whose sizes are from nano scale to micro scale. Therefore, microbes can be used as natural "building blocks" in nano/micro multi-level fabrication processes. At present, most of the bio-manufacturing methods do not apply to direct control of living microbes. Their microbiological global functions and superiorities are not available. In this paper, two novel nano/micro bio-fabrication approaches, micro-fluidic control method and magnetic control method have been established. The living microbes could be manipulated to form micro-scaled patterns or to move orientedly. By these approaches, living microbes are taken as nano/micro robots. We could employ their specific biological functions and regulate their controllable self-assembly, which is expected to design and create a series of new special functional materials and devices.

Optimal Supply of Precursors for CoQ10 Production by Rhodopseudomonas Palustris / 现代生物医学进展

CoQ10 has been used not only as a medicine but also as food supplements because of its various physiological and biochemical activities. A full-factorial central composite design and response surface methodology were used for optimizing three precursors Solanesol, 4-hydroxybenzoic acid and methionine to maximize the production of CoQ10 by Rhodopseudomonas palustris J001. The optimization of the model predicted a maximum response 40.6 [(mg CoQ10)(g dried biomass)-1] CoQ10 production with 124.8 mg l-1 Solanesol, 267.7 mg l-14-hydroxybenzoic acid and 130.2 mg l-1 methionine, respectively. A new combination was prepared according to the result. The observed response was 40.5 ± 0.2 [(mg CoQ10)(g dried biomass)-1] and was 109.8%higher than in the control with no addition of the three precursors.

Kinetic model for optimal feeding strategy in astaxanthin production by Xanthophyllomyces dendrorhous / 生物工程学报

Astaxanthin is a useful pigmentation source in fish aquaculture. It has strong antioxidative activity and therefore has potential application in delaying aging and degenerative diseases in human and animals. In recent years, there is a growing demand for astaxanthin. The red yeast Xanthophyllomyces dendrorhous (called Phaffia rhodozyma before) is one of the most promising microorganisms for the commercial production of astaxanthin. During fermentation, X. dendrorhous shows the Crabtree effect. Higher glucose concentration will cause significant reductions in biomass and astaxanthin production. Therefore, fed-batch processes are particularly useful. In this paper, effects of glucose-feeding strategies on astaxanthin production by X. dendrorhous were studied. Based on the substrate inhibition model, an optimized two-stage feeding strategy for astaxanthin production of high-cell-density fermentation was proposed. Glucose concentration was first controlled at about 25 g/L during the lag phase and the early exponential phase. In such case, biomass could reach its maximum value in relatively short time. Then the glucose concentration was controlled at about 5 g/L in the later exponential phase and stationary phase. The synthesis of astaxanthin could be effectively prolonged. The results showed that the optimized two-stage feeding strategy was the best among all the feeding strategies, and could obtain the highest biomass (23.8 g/L) and astaxanthin production (29.05 mg/L), which was a significant increase (52.8% and 109% respectively) compared with a batch process.

Culture Conditions Optimization of Magnetotactic Bacteria WM-1 / 现代生物医学进展

The culture of Magnetospirillum magneticum WM-1 depends on several control factors that have great effect on the magnetic cells concentration. Investigation into the optimal culture conditions needs a large number of experiments. So it is desirable to minimize the number of experiments and maximize the information gained from them. The orthogonal design of experiments and mathematical statistical method are considered as effective methods to optimize the culture condition of magnetotactic bacteria WM-1 for high magnetic cells concentration. The effects of the four factors, such as pH value of medium, oxygen concentration of gas phase in the serum bottle, C:C (mtartaric acid: msuccinic acid) ratio and NaNO3 concentration, are simultaneously investigated by only sixteen experiments through the orthogonal design L16(44) method. The optimal culture condition is obtained. At the optimal culture condition (pH 7.0, a oxygen concentration 4.0%, C: C (mtartaric acid: msuccinic acid) ratio 1:2 and NaNO3 100 mg l-1), the magnetic cells concentration is promoted to 6.5×107 cells ml-1, approximately 8.3% higher than that under the initial conditions. The pH value of medium is very important factor for magnetic cells concentration. It can be proved that the orthogonal design of experiment is of 90% confidence. The results from the hysteresis of WM-1 shows that Hc = 230 Oe, Ms = 0.9 emu/g dry wt. Cells,and Mr / Ms = 0.50.