低能N+注入介导构建丙三醇高产多形汉逊酵母重组菌株
钱卫东 1赵德志 1宁肖肖 1陈雪峰 1蔡长龙 2毛培宏3
作者信息
- 1. 陕西科技大学生命科学与工程学院，陕西 西安 710021
- 2. 西安工业大学光电学院离子束生物工程研究中心，陕西 西安 710032
- 3. 新疆大学物理科学与技术学院离子束生物技术中心，新疆 乌鲁木齐 830008
- 折叠
摘要
目的：获得丙三醇高产多形汉逊酵母重组菌株。方法：利用低能氮离子（N+）介导转基因技术介导多形汉逊酵母基因组总DNA随机转化自身，构建丙三醇高产工程菌。通过高盐培养基初筛，纸色谱法和气相色谱法复筛目标菌株。进而运用正交试验对发酵培养基进行优化。结果：获得一株遗传稳定的能够高效生产丙三醇的多形汉逊酵母重组菌株，其产量高达70.29g/L。对重组菌株传代培养8代，利用气相色谱分析发现丙三醇产量基本稳定。结论：利用低能N+离子介导多形汉逊酵母基因组转化自身宿主技术，可以在丙三醇生物合成及相关调控基因未知的情况下构建丙三醇高产的多形汉逊酵母重组菌株。
Abstract
Objective:To obtain recombinant Hansenula polymorpha strain with high yield of glycerol. Methods:We have developed an efficient method to construct recombinant Hansenula polymorpha strains with enhanced ability to produce glycerol by randomly incorporating total genomic DNA of Hansenula polymorpha into itself through low-energy nitrogen ions (N+)-mediated genetic transformation. To screen efficiently target recombinant strains, a tolerant to high salt culture-based high-throughout screening was utilized. Then paper chromatography and gas chromatography were used to further analyze the candidate strains based on the preliminary screening. Additional y, the orthogonal experiment was used to optimize the fermentation medium. Results:One strain of genetical y stable recombinant Hansenula polymorpha strain capable of yielding efficiently glycerol was constructed, and its yield is up to 70.29g/L. Meanwhile, the recombinant strain was subcultured for eight generations, and the output is stable based on the GC analysis. Conclusion:Hansenula polymorpha total genomic DNA can be transformed randomly into itself using low-energy nitrogen ions (N+)-mediated genetic transformation to obtain recombinant Hansenula polymorpha strains with high-yield of glycerol when the biosynthesis of glycerol is stil unknown .
关键词
低能N+离子注入/遗传转化/多形汉逊酵母Key words
low-energy nitrogen ions (N+)-mediated/genetic transformation/Hansenula polymorpha, glycerol/high-yield引用本文复制引用
基金项目
国家自然科学基金(31100040)
陕西省教育厅项目(2013JK0727)
西安市未央区项目(201210)
出版年
2014
NETL
NSTL
万方数据