To avoid the long time required for conventional sexual crossing, transgenic tobacco (Nicotiana tabacum L.) plants harboring phbA gene (encoding 3_ketothiolase) were used as the target plant for the second transfo...To avoid the long time required for conventional sexual crossing, transgenic tobacco (Nicotiana tabacum L.) plants harboring phbA gene (encoding 3_ketothiolase) were used as the target plant for the second transformation mediated by Agrobacterium tumefaciens (Smith et Townsend) Conn LBA4404 containing pZCB which was constructed by linking phbB (encoding acetoacetyl_CoA reductase), phbC (encoding PHB synthase) and ctp sequence to pBIB_HYG under the control of CaMV 35S promoter. The hygromycin resistant transformants were morphologically normal and stable integration of phbB and phbC was confirmed by PCR and PCR_Southern. Moreover, RT_PCR_DNA hybridization analysis showed that 6.67% of the transformed tobacco plants could express both phbB and phbC at transcriptional level.展开更多
The gene of the first key enzyme of poly-β-hydroxybutyrate synthesis, 3-ketothiolase, has been amplified and cloned from chromosomal DNA of Alcaligenes eutrophus H16 by PCR. DNA sequencing results show that phbA clon...The gene of the first key enzyme of poly-β-hydroxybutyrate synthesis, 3-ketothiolase, has been amplified and cloned from chromosomal DNA of Alcaligenes eutrophus H16 by PCR. DNA sequencing results show that phbA cloned in pBluescriptSK^+ has an identical sequence with that reported previously except for one base pair. The plant constitutive expression vector has been constructed and tobacco has been transformed in order to examine the phbA gene function and the efficiency of ctp gene product. SDS-polyacrylamide gel electrophoresis result shows that the ctp gene product could direct foreign protein into plastid efficiently and phbA gene could be translated into corresponding protein with correct size. The enzyme activity analysis of 3-ketothiolase shows that the enzyme could catalyze acetyl-CoA to form acetoacetylCoA.展开更多
以钴铜为主催化剂,镍锆为助催化剂,TiO_2为载体,用浸渍法制备一系列负载型催化剂,并用XRD、IR对其进行表征。将催化剂用于对甲酚常压液相催化氧化制备对羟基苯甲醛(PHBA)的反应中,结果表明,n(Co^(2+))∶n(Cu^(2+))=3∶1,n(Co^(2+))∶n(N...以钴铜为主催化剂,镍锆为助催化剂,TiO_2为载体,用浸渍法制备一系列负载型催化剂,并用XRD、IR对其进行表征。将催化剂用于对甲酚常压液相催化氧化制备对羟基苯甲醛(PHBA)的反应中,结果表明,n(Co^(2+))∶n(Cu^(2+))=3∶1,n(Co^(2+))∶n(Ni^(2+))=1∶0.06,n(Co^(2+))∶n(Zr^(2+))=1∶0.005时,催化剂的催化效果最佳。同时,考察了反应温度、氢氧化钠用量、反应时间、通氧量、甲醇用量对该反应的影响。在单因素实验基础上,采用正交实验优化出该反应较佳工艺:反应时间为5 h,反应温度为75℃,甲醇用量为30 m L,n(氢氧化钠)∶n(对甲酚)=5∶1,通氧量为50 m L/min。以对甲酚计,PHBA的收率为92.58%。所制备催化剂在常压下对于对甲酚的催化氧化反应具有较高的催化活性和重复性。展开更多
Coenzyme Q10 (CoQ10), an important antioxidant molecule playing a major role in electron transport chain, has been commercially produced by fermentation process for the use in oral nutraceutical formulations. Construc...Coenzyme Q10 (CoQ10), an important antioxidant molecule playing a major role in electron transport chain, has been commercially produced by fermentation process for the use in oral nutraceutical formulations. Constructing the high-yielding CoQ10 producing strains is a pre-requisite for cost-effective production. A superior mutant strain P-87 generated from Paracoccus denitrificans ATCC 19367, which showed 1.25-fold improvement in specific CoQ10 content higher than the wild type strain at shake flask level, was selected to carry out the studies on CoQ10 yield improvement through fermenter process optimization. In the course of study, initially the cane-molasses-based medium and fed-batch fermentation strategy using pHBA in combination with sucrose were standardized in shake flask using wild type strain. This strategy was subsequently translated at 2 L laboratory fermenter while optimizing the fermentation process parameters using improved mutant strain P-87. Under optimized fermentation condition, mutant strain P-87 produced 49.85 mg/L of CoQ10 having specific content of 1.63 mg/g of DCW, which was 1.36 folds higher than the specific CoQ10 content of wild-type strain under similar optimized condition. The temperature and DO were found to be critical parameters for CoQ10 production by mutant strain P-87. The optimum temperature was found to be 32°C and the optimum DO concentration to be maintained throughout the fermentation cycle was found to be 30% of air saturation. Overall, a new cost-effective process has been established for the production of CoQ10 using the cheaper substrate “cane molasses” and higher CoQ10 producing mutant strain P-87.展开更多
Two models of PHBA adsorbed on the surface of silver nanoparticles were established, and each of them corresponding to an experimental configuration: the first model is PHBA adsorbed on one Ag atom through carboxyl, a...Two models of PHBA adsorbed on the surface of silver nanoparticles were established, and each of them corresponding to an experimental configuration: the first model is PHBA adsorbed on one Ag atom through carboxyl, and the second one is PHBA adsorbed on two Ag atoms through carboxyl and hydroxyl. And the Raman spectra of these two models using DFT-B3PW91 with lanl2dz were calculated; it was found that the calculated Raman frequencies are in good agreement with experimental values. So one can conclude that the simplified models are probably reasonable to describe some surface enhanced Raman experiments.展开更多
文摘To avoid the long time required for conventional sexual crossing, transgenic tobacco (Nicotiana tabacum L.) plants harboring phbA gene (encoding 3_ketothiolase) were used as the target plant for the second transformation mediated by Agrobacterium tumefaciens (Smith et Townsend) Conn LBA4404 containing pZCB which was constructed by linking phbB (encoding acetoacetyl_CoA reductase), phbC (encoding PHB synthase) and ctp sequence to pBIB_HYG under the control of CaMV 35S promoter. The hygromycin resistant transformants were morphologically normal and stable integration of phbB and phbC was confirmed by PCR and PCR_Southern. Moreover, RT_PCR_DNA hybridization analysis showed that 6.67% of the transformed tobacco plants could express both phbB and phbC at transcriptional level.
文摘The gene of the first key enzyme of poly-β-hydroxybutyrate synthesis, 3-ketothiolase, has been amplified and cloned from chromosomal DNA of Alcaligenes eutrophus H16 by PCR. DNA sequencing results show that phbA cloned in pBluescriptSK^+ has an identical sequence with that reported previously except for one base pair. The plant constitutive expression vector has been constructed and tobacco has been transformed in order to examine the phbA gene function and the efficiency of ctp gene product. SDS-polyacrylamide gel electrophoresis result shows that the ctp gene product could direct foreign protein into plastid efficiently and phbA gene could be translated into corresponding protein with correct size. The enzyme activity analysis of 3-ketothiolase shows that the enzyme could catalyze acetyl-CoA to form acetoacetylCoA.
文摘以钴铜为主催化剂,镍锆为助催化剂,TiO_2为载体,用浸渍法制备一系列负载型催化剂,并用XRD、IR对其进行表征。将催化剂用于对甲酚常压液相催化氧化制备对羟基苯甲醛(PHBA)的反应中,结果表明,n(Co^(2+))∶n(Cu^(2+))=3∶1,n(Co^(2+))∶n(Ni^(2+))=1∶0.06,n(Co^(2+))∶n(Zr^(2+))=1∶0.005时,催化剂的催化效果最佳。同时,考察了反应温度、氢氧化钠用量、反应时间、通氧量、甲醇用量对该反应的影响。在单因素实验基础上,采用正交实验优化出该反应较佳工艺:反应时间为5 h,反应温度为75℃,甲醇用量为30 m L,n(氢氧化钠)∶n(对甲酚)=5∶1,通氧量为50 m L/min。以对甲酚计,PHBA的收率为92.58%。所制备催化剂在常压下对于对甲酚的催化氧化反应具有较高的催化活性和重复性。
文摘Coenzyme Q10 (CoQ10), an important antioxidant molecule playing a major role in electron transport chain, has been commercially produced by fermentation process for the use in oral nutraceutical formulations. Constructing the high-yielding CoQ10 producing strains is a pre-requisite for cost-effective production. A superior mutant strain P-87 generated from Paracoccus denitrificans ATCC 19367, which showed 1.25-fold improvement in specific CoQ10 content higher than the wild type strain at shake flask level, was selected to carry out the studies on CoQ10 yield improvement through fermenter process optimization. In the course of study, initially the cane-molasses-based medium and fed-batch fermentation strategy using pHBA in combination with sucrose were standardized in shake flask using wild type strain. This strategy was subsequently translated at 2 L laboratory fermenter while optimizing the fermentation process parameters using improved mutant strain P-87. Under optimized fermentation condition, mutant strain P-87 produced 49.85 mg/L of CoQ10 having specific content of 1.63 mg/g of DCW, which was 1.36 folds higher than the specific CoQ10 content of wild-type strain under similar optimized condition. The temperature and DO were found to be critical parameters for CoQ10 production by mutant strain P-87. The optimum temperature was found to be 32°C and the optimum DO concentration to be maintained throughout the fermentation cycle was found to be 30% of air saturation. Overall, a new cost-effective process has been established for the production of CoQ10 using the cheaper substrate “cane molasses” and higher CoQ10 producing mutant strain P-87.
文摘Two models of PHBA adsorbed on the surface of silver nanoparticles were established, and each of them corresponding to an experimental configuration: the first model is PHBA adsorbed on one Ag atom through carboxyl, and the second one is PHBA adsorbed on two Ag atoms through carboxyl and hydroxyl. And the Raman spectra of these two models using DFT-B3PW91 with lanl2dz were calculated; it was found that the calculated Raman frequencies are in good agreement with experimental values. So one can conclude that the simplified models are probably reasonable to describe some surface enhanced Raman experiments.
