通过简单加热氯代正丁基吡啶([C_4Pyr]Cl)和对甲苯磺酸(TsOH)的混合物制备了[C_4Pyr]Cl/nTsOH,(n=0.1,0.2,0.3)型低共熔溶剂。以[C_4Pyr]Cl/nTsOH为催化剂和萃取剂,H_2O_2为氧化剂组成萃取-催化氧化脱硫体系氧化脱除模拟油中的硫化物。...通过简单加热氯代正丁基吡啶([C_4Pyr]Cl)和对甲苯磺酸(TsOH)的混合物制备了[C_4Pyr]Cl/nTsOH,(n=0.1,0.2,0.3)型低共熔溶剂。以[C_4Pyr]Cl/nTsOH为催化剂和萃取剂,H_2O_2为氧化剂组成萃取-催化氧化脱硫体系氧化脱除模拟油中的硫化物。通过FTIR表征,确定[C_4Pyr]Cl/0.2TsOH的结构以及氧化产物,并考察了不同脱硫体系、n(TsOH)∶n([C_4Pyr]Cl)、低共熔溶剂加入量、反应温度、n(H_2O_2)∶n(二苯并噻吩)和含硫化物类型对脱硫效果的影响。实验结果表明,在低共熔溶剂[C_4Pyr]Cl/0.2TsOH加入量1.00 m L、反应温度50℃、n(H_2O_2)∶n(二苯并噻吩)=6、模拟油用量5 m L的反应条件下,[C_4Pyr]Cl/0.2TsOH对二苯并噻吩、4,6-二甲基二苯并噻吩和苯并噻吩的脱硫率分别达98.2%,96.0%,40.2%。由一级动力学方程和Arrhenius方程计算氧化脱除二苯并噻吩所需的表观活化能约为51.95 k J/mol。[C_4Pyr]Cl/0.2TsOH回收利用5次后,脱硫率仍不低于95.1%。展开更多
Gibberellic acid(GA3)is a vital plant growth hormone widely used in agriculture.Currently,GA3 production relies on liquid fermentation by the filamentous fungus Fusarium fujikuroi.However,the lack of an effective sele...Gibberellic acid(GA3)is a vital plant growth hormone widely used in agriculture.Currently,GA3 production relies on liquid fermentation by the filamentous fungus Fusarium fujikuroi.However,the lack of an effective selection marker recycling system hampers the application of metabolic engineering technology in F.fujikuroi,as multiple-gene editing and positive-strain screening still rely on a limited number of antibiotics.In this study,we developed a strategy using pyr4-blaster and CRISPR/Cas9 tools for recycling orotidine-5'-phosphate decarboxylase(Pyr4)selection markers.We demonstrated the effectiveness of this method for iterative gene integration and large gene-cluster deletion.We also successfully improved GA3 titers by overexpressing geranylgeranyl pyrophosphate synthase and truncated 3-hydroxy-3-methyl glutaryl coenzyme A reductase,which rewired the GA3 biosynthesis pathway.These results highlight the efficiency of our established system in recycling selection markers during iterative gene editing events.Moreover,the selection marker recycling system lays the foundation for further research on metabolic engineering for GA3 industrial production.展开更多
文摘通过简单加热氯代正丁基吡啶([C_4Pyr]Cl)和对甲苯磺酸(TsOH)的混合物制备了[C_4Pyr]Cl/nTsOH,(n=0.1,0.2,0.3)型低共熔溶剂。以[C_4Pyr]Cl/nTsOH为催化剂和萃取剂,H_2O_2为氧化剂组成萃取-催化氧化脱硫体系氧化脱除模拟油中的硫化物。通过FTIR表征,确定[C_4Pyr]Cl/0.2TsOH的结构以及氧化产物,并考察了不同脱硫体系、n(TsOH)∶n([C_4Pyr]Cl)、低共熔溶剂加入量、反应温度、n(H_2O_2)∶n(二苯并噻吩)和含硫化物类型对脱硫效果的影响。实验结果表明,在低共熔溶剂[C_4Pyr]Cl/0.2TsOH加入量1.00 m L、反应温度50℃、n(H_2O_2)∶n(二苯并噻吩)=6、模拟油用量5 m L的反应条件下,[C_4Pyr]Cl/0.2TsOH对二苯并噻吩、4,6-二甲基二苯并噻吩和苯并噻吩的脱硫率分别达98.2%,96.0%,40.2%。由一级动力学方程和Arrhenius方程计算氧化脱除二苯并噻吩所需的表观活化能约为51.95 k J/mol。[C_4Pyr]Cl/0.2TsOH回收利用5次后,脱硫率仍不低于95.1%。
基金supported by National key research and development program of China(2021YFC2104300)Natural Science Foundation of Jiangsu Province(BK20210573,20230381)+2 种基金Project of Leading Innovative Talents in Short-term Jiangxi Province(jxsq2023102172)Tianjin Synthetic Biotechnology Innovation Capacity Improvement Project TSBICIP-PTJS-003-04Postgraduate Research&Practice Innovation Program of Jiangsu Province KYCX23_1788.
文摘Gibberellic acid(GA3)is a vital plant growth hormone widely used in agriculture.Currently,GA3 production relies on liquid fermentation by the filamentous fungus Fusarium fujikuroi.However,the lack of an effective selection marker recycling system hampers the application of metabolic engineering technology in F.fujikuroi,as multiple-gene editing and positive-strain screening still rely on a limited number of antibiotics.In this study,we developed a strategy using pyr4-blaster and CRISPR/Cas9 tools for recycling orotidine-5'-phosphate decarboxylase(Pyr4)selection markers.We demonstrated the effectiveness of this method for iterative gene integration and large gene-cluster deletion.We also successfully improved GA3 titers by overexpressing geranylgeranyl pyrophosphate synthase and truncated 3-hydroxy-3-methyl glutaryl coenzyme A reductase,which rewired the GA3 biosynthesis pathway.These results highlight the efficiency of our established system in recycling selection markers during iterative gene editing events.Moreover,the selection marker recycling system lays the foundation for further research on metabolic engineering for GA3 industrial production.
