Synthetic Biology Journal ›› 2023, Vol. 4 ›› Issue (5): 1000-1019.DOI: 10.12211/2096-8280.2023-031
• Invited Review • Previous Articles Next Articles
Yujie WU1,2,3, Xinxin LIU1, Jianhui LIU1, Kaiguang Yang1, Zhigang SUI1, Lihua ZHANG1, Yukui ZHANG1
Received:
2023-04-17
Revised:
2023-06-26
Online:
2023-11-15
Published:
2023-10-31
Contact:
Kaiguang Yang, Zhigang SUI
吴玉洁1,2,3, 刘欣欣1, 刘健慧1, 杨开广1, 随志刚1, 张丽华1, 张玉奎1
通讯作者:
杨开广,随志刚
作者简介:
基金资助:
CLC Number:
Yujie WU, Xinxin LIU, Jianhui LIU, Kaiguang Yang, Zhigang SUI, Lihua ZHANG, Yukui ZHANG. Research progress of strain screening and quantitative analysis of key molecules based on high-throughput liquid chromatography and mass spectrometry[J]. Synthetic Biology Journal, 2023, 4(5): 1000-1019.
吴玉洁, 刘欣欣, 刘健慧, 杨开广, 随志刚, 张丽华, 张玉奎. 基于高通量液相色谱质谱技术的菌株筛选与关键分子定量分析研究进展[J]. 合成生物学, 2023, 4(5): 1000-1019.
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URL: https://synbioj.cip.com.cn/EN/10.12211/2096-8280.2023-031
质谱 技术 | 优缺点 | 菌株培养 方式 | 筛选通量 | 适用对象 | 应用示例 | 参考 文献 |
---|---|---|---|---|---|---|
MALDI | 优点:高耐盐性,易于操作,样品用量少,分析速度快,通量高 缺点:需要添加基质,可能引入基质峰干扰 | 琼脂平板 | 约2 s/样品 | 可用于分析脂肪酸等小分子及蛋白质等生物大分子 | 通过检测酰基链磷脂酰胆碱,以筛选高产中链脂肪酸的酿酒酵母菌株 | [ |
琼脂平板 | <2.5 s/样品 | 机器视觉算法辅助识别随机分布菌落,应用于大肠杆菌天然产物文库筛选 | [ | |||
琼脂平板/ 微孔板 | <5 s/样品 | 开发了菌落位置转换MALDI坐标算法,用于酿酒酵母、大肠杆菌及荧光假单胞菌文库筛选 | [ | |||
微孔板 | 约5 s/样品 | 筛选大肠杆菌环二肽合酶突变体文库及羊毛硫肽化合物文库,基本实现整个流程自动化 | [ | |||
液滴 | 约5 s/样品 | 酵母甜蛋白及植酸酶产物文库筛选 | [ | |||
SAMDI | 优点:特异性强,可用于检测复杂体系中的目标分子 缺点:无法区分分子量相同的产物和底物 | 琼脂平板 | <1 s/样品 | 可用于分析小分子 | 利用自组转单层技术在靶板上修饰马来酰亚胺基团,筛选大肠杆菌细胞色素P411突变体文库 | [ |
NIMS | 优点:灵敏度高,无需添加基质 缺点:需要构建纳米结构表面 | 琼脂平板 | <1 s/样品 | 可用于分析小分子、脂质和肽 | 结合点击化学,筛选大肠杆菌细胞色素P450突变体文库 | [ |
ESI | 优点:采用软电离方式,基本不产生碎片峰,分辨率高 缺点:耐盐性较差,取样效率低 | 微孔板 | 约84 s/样品 | 可用于分析极性强、热稳定性差的小分子和生物大分子 | 液相色谱质谱联用快速筛选三萜桦木酸产量提高的酿酒酵母菌株 | [ |
液滴 | 约1 s/样品 | 在线筛选谷氨酸棒杆菌文库 | [ | |||
微孔板 | 约60 s/样品 | 融合LESA技术,用于酵母菌株衣康酸、三乙酸内酯和棕榈酸产物定量筛选分析 | [ | |||
DESI | 优点:高耐盐性,可实现原位实时分析,通量高 缺点:电喷雾溶剂组成可能影响样品溶解与电离 | 琼脂平板 | <1 s/样品 | 可用于分析非极性小分子及极性大分子 | 与成像技术结合,快速筛选大肠杆菌突变体文库 | [ |
LA-REI | 优点:易于操作,无需样品制备,快速分析 缺点:无法区分异构体 | 琼脂平板 | 约10 s/样品 | 可用于分析小分子代谢物 | 激光辅助实现酵母白桦酸产物文库筛选 | [ |
Table 1 Summary of strain screening studies based on high-throughput mass spectrometry
质谱 技术 | 优缺点 | 菌株培养 方式 | 筛选通量 | 适用对象 | 应用示例 | 参考 文献 |
---|---|---|---|---|---|---|
MALDI | 优点:高耐盐性,易于操作,样品用量少,分析速度快,通量高 缺点:需要添加基质,可能引入基质峰干扰 | 琼脂平板 | 约2 s/样品 | 可用于分析脂肪酸等小分子及蛋白质等生物大分子 | 通过检测酰基链磷脂酰胆碱,以筛选高产中链脂肪酸的酿酒酵母菌株 | [ |
琼脂平板 | <2.5 s/样品 | 机器视觉算法辅助识别随机分布菌落,应用于大肠杆菌天然产物文库筛选 | [ | |||
琼脂平板/ 微孔板 | <5 s/样品 | 开发了菌落位置转换MALDI坐标算法,用于酿酒酵母、大肠杆菌及荧光假单胞菌文库筛选 | [ | |||
微孔板 | 约5 s/样品 | 筛选大肠杆菌环二肽合酶突变体文库及羊毛硫肽化合物文库,基本实现整个流程自动化 | [ | |||
液滴 | 约5 s/样品 | 酵母甜蛋白及植酸酶产物文库筛选 | [ | |||
SAMDI | 优点:特异性强,可用于检测复杂体系中的目标分子 缺点:无法区分分子量相同的产物和底物 | 琼脂平板 | <1 s/样品 | 可用于分析小分子 | 利用自组转单层技术在靶板上修饰马来酰亚胺基团,筛选大肠杆菌细胞色素P411突变体文库 | [ |
NIMS | 优点:灵敏度高,无需添加基质 缺点:需要构建纳米结构表面 | 琼脂平板 | <1 s/样品 | 可用于分析小分子、脂质和肽 | 结合点击化学,筛选大肠杆菌细胞色素P450突变体文库 | [ |
ESI | 优点:采用软电离方式,基本不产生碎片峰,分辨率高 缺点:耐盐性较差,取样效率低 | 微孔板 | 约84 s/样品 | 可用于分析极性强、热稳定性差的小分子和生物大分子 | 液相色谱质谱联用快速筛选三萜桦木酸产量提高的酿酒酵母菌株 | [ |
液滴 | 约1 s/样品 | 在线筛选谷氨酸棒杆菌文库 | [ | |||
微孔板 | 约60 s/样品 | 融合LESA技术,用于酵母菌株衣康酸、三乙酸内酯和棕榈酸产物定量筛选分析 | [ | |||
DESI | 优点:高耐盐性,可实现原位实时分析,通量高 缺点:电喷雾溶剂组成可能影响样品溶解与电离 | 琼脂平板 | <1 s/样品 | 可用于分析非极性小分子及极性大分子 | 与成像技术结合,快速筛选大肠杆菌突变体文库 | [ |
LA-REI | 优点:易于操作,无需样品制备,快速分析 缺点:无法区分异构体 | 琼脂平板 | 约10 s/样品 | 可用于分析小分子代谢物 | 激光辅助实现酵母白桦酸产物文库筛选 | [ |
质谱采集类型 | 菌株样本 | 分析通量(每天检测样本数估值) | 参考文献 |
---|---|---|---|
靶向采集 | 大肠杆菌E.coil | 25 min分离梯度定量大肠杆菌中600多条肽(约50样品/天) | [ |
非靶向采集 | 酿酒酵母S.cerevistae | 19 min有效分离梯度定量检测796个酵母菌株蛋白质组(约70样品/天) | [ |
非靶向采集 | 大肠杆菌E.coil | 4 min分离梯度定量226个脂类分子(约320样品/天) | [ |
非靶向采集 | 酿酒酵母S.cerevistae | 15 min分离梯度结合双分析柱系统检测蛋白(约90样品/天) | [ |
非靶向采集 | 酿酒酵母S.cerevistae | 23 min分离梯度定量检测100个酵母蛋白质组(约45样品/天) | [ |
非靶向采集 | 酿酒酵母S.cerevistae | 5 min分离梯度定量1900种蛋白(约280样品/天) | [ |
靶向采集 | 大肠杆菌E.coil | 10 min分离梯度定量400多种蛋白质(约130样品/天) | [ |
靶向采集 | 恶臭假单胞菌P.putida KT2440 | 32 min分离梯度定量132种蛋白质(约40样品/天) | [ |
靶向采集 | 大肠杆菌E.coil | 5 min分离梯度检测102种代谢物(约280样品/天) | [ |
Table 2 Summary of quantitative studies of key molecules of strains based on liquid chromatography mass spectrometry
质谱采集类型 | 菌株样本 | 分析通量(每天检测样本数估值) | 参考文献 |
---|---|---|---|
靶向采集 | 大肠杆菌E.coil | 25 min分离梯度定量大肠杆菌中600多条肽(约50样品/天) | [ |
非靶向采集 | 酿酒酵母S.cerevistae | 19 min有效分离梯度定量检测796个酵母菌株蛋白质组(约70样品/天) | [ |
非靶向采集 | 大肠杆菌E.coil | 4 min分离梯度定量226个脂类分子(约320样品/天) | [ |
非靶向采集 | 酿酒酵母S.cerevistae | 15 min分离梯度结合双分析柱系统检测蛋白(约90样品/天) | [ |
非靶向采集 | 酿酒酵母S.cerevistae | 23 min分离梯度定量检测100个酵母蛋白质组(约45样品/天) | [ |
非靶向采集 | 酿酒酵母S.cerevistae | 5 min分离梯度定量1900种蛋白(约280样品/天) | [ |
靶向采集 | 大肠杆菌E.coil | 10 min分离梯度定量400多种蛋白质(约130样品/天) | [ |
靶向采集 | 恶臭假单胞菌P.putida KT2440 | 32 min分离梯度定量132种蛋白质(约40样品/天) | [ |
靶向采集 | 大肠杆菌E.coil | 5 min分离梯度检测102种代谢物(约280样品/天) | [ |
软件工具 | 分析对象 | 网址 | 参考文献 |
---|---|---|---|
AB3D | 蛋白质组 | http://www.first-ms3d.jp/english/(Mass++插件) | [ |
DynaMet | 代谢组 | https://pypi.python.org/pypi/dynamet/ | [ |
DIA-NN | 蛋白质组 | https://github.com/vdemichev/diann | [ |
Tric | 蛋白质组 | http://proteomics.ethz.ch/tric/ | [ |
MRMAnalyzer | 代谢组 | http://link.springer.com/10.1007/s11306-015-0809-4(R包) | [ |
MESSI | 代谢组 | http://sbb.hku.hk/MESSI/ | [ |
PTools | 代谢组 | https://brg.ai.sri.com/ptools/ | [ |
Table 3 Summary of data processing software and bioinformatics tools in the quantitative analysis of key molecules
软件工具 | 分析对象 | 网址 | 参考文献 |
---|---|---|---|
AB3D | 蛋白质组 | http://www.first-ms3d.jp/english/(Mass++插件) | [ |
DynaMet | 代谢组 | https://pypi.python.org/pypi/dynamet/ | [ |
DIA-NN | 蛋白质组 | https://github.com/vdemichev/diann | [ |
Tric | 蛋白质组 | http://proteomics.ethz.ch/tric/ | [ |
MRMAnalyzer | 代谢组 | http://link.springer.com/10.1007/s11306-015-0809-4(R包) | [ |
MESSI | 代谢组 | http://sbb.hku.hk/MESSI/ | [ |
PTools | 代谢组 | https://brg.ai.sri.com/ptools/ | [ |
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