Acta Horticulturae Sinica ›› 2024, Vol. 51 ›› Issue (7): 1516-1528.doi: 10.16420/j.issn.0513-353x.2023-0528
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LI Xin1,*(), CHAI Yingfang2, TIAN Zhen2, WANG Pengwei2, CHENG Yunjiang2
Received:
2023-09-27
Revised:
2024-05-05
Online:
2024-07-25
Published:
2024-07-19
Contact:
LI Xin
LI Xin, CHAI Yingfang, TIAN Zhen, WANG Pengwei, CHENG Yunjiang. Isolation and Purification of Mitochondria and Its Application in Research on Fruit Ripening and Senescence[J]. Acta Horticulturae Sinica, 2024, 51(7): 1516-1528.
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URL: https://www.ahs.ac.cn/EN/10.16420/j.issn.0513-353x.2023-0528
种类 Category | 果实 Fruit | 组织 Tissue | 分离方法 Method | 参考文献 Reference |
---|---|---|---|---|
仁果类 Pome fruit | 苹果 Apple | 果肉Pulp | DC (1R) | Lieberman, |
果肉Pulp | DC (1R) + W (1T) | Neal & Hulme, Lieberman, | ||
果皮Peel | DC (1R) + W (1T) | Hulme et al., | ||
果肉Pulp | DC (2R) | Romani et al., | ||
果肉Pulp | DC (2R) + DPGC + SPGC + W (2T) | Qin et al., | ||
梨 Pear | 果肉Pulp | DC (2R) | Romani et al., | |
DC (1R) + W (1T) + CSGC | Miller & Romani, | |||
DC (1R) + W (1T) | Li et al., | |||
核果类 Stone fruit | 鳄梨 Avocado | 果肉Pulp | DC (1R) + W (1T) | Biale et al., |
DC (3R) | Wiskich et al., | |||
DC (2R) | Lance et al., | |||
DC (1R) + W (1T) + SPGC + W (2T) | Moreau & Romani, | |||
芒果 Mango | 果肉Pulp | DC (2R) | Baqui et al., | |
DC (1R) + W (1T) | Kane & Marcellin, | |||
桃 Peach | 果肉Pulp | DC (1R) + W (1T) | Kan et al., | |
DC (1R) + W (2T) | Jin et al., | |||
DC (1R) + DPGC + W (1T) + SPGC + W (2T) | Qin et al., | |||
DC (1R) + DPGC + W (2T) | Yang et al., | |||
DC (1R) + W (1T) + DSGC + W (1T) | Jing et al., | |||
DC (1R) + DPGC + W (2T) | Wu et al., | |||
DC (1R) + DSGC + W (1T) | Wang et al., | |||
浆果类 Berries | 番茄 Tomato | 果肉Pulp | DC (1R) + W (1T) | Dickinson & Hanson, |
DC (2R) | Ku et al., | |||
DC (1R) + W (2T) | Li et al., | |||
DC (1R) + W (1T) + CSGC | Kalra & Brooks, | |||
DC (2R) + DPGC | Jeffery et al., | |||
DC (1R) + W (2T) + SPGC + W (2T) | Jarmuszkiewicz et al., | |||
DC (1R) + SPGC + W (2T) | Holtzapffel et al., | |||
DC (2R) + DPGC + W (2T) | de Oliveira et al., | |||
DC (1R) + DPGC + W (2T) + SPGC + W (2T) | Cai et al., | |||
香蕉 Banana | 果肉Pulp | DC (2R) | Haard & Hultin, | |
果皮Peel | DC (1R) + W (2T) + CSGC + W (1T) | Chotikakham et al., | ||
蓝莓 Blueberry | 果肉Pulp | DC (1R) + W (1T) | Zhou et al., | |
番木瓜Papaya 草莓Strawberry 番石榴Guava | 果肉Pulp | DC (2R) + DPGC + W (2T) | de Oliveira et al., | |
猕猴桃Kiwifruit | 果肉Pulp | DC (1R) + W (2T) | Ali et al., | |
菠萝 Pineapple | 果肉Pulp | DC (1R) + DSGC | Nukuntornprakit et al., | |
柑果类 Citrus | 柠檬Lemon | 果肉Pulp | DC (1R) + W (1T) | Bogin & Erickson, |
甜橙Orange | DC (2R) | Vines & Oberbacher, | ||
甜橙Orange | DC (1R) | Katz et al., | ||
柑Tangerine 橘Mandarin 橙Orange 柚Pummelo | DC (2R) + DPGC + W (2T) | Li et al., | ||
瓜类 Melon | 甜瓜 Muskmelon | 果肉Pulp | DC (2R) + DPGC + SPGC + W (2T) | Lyu et al., |
Table 1 Summary for isolation and purification of fruit mitochondria
种类 Category | 果实 Fruit | 组织 Tissue | 分离方法 Method | 参考文献 Reference |
---|---|---|---|---|
仁果类 Pome fruit | 苹果 Apple | 果肉Pulp | DC (1R) | Lieberman, |
果肉Pulp | DC (1R) + W (1T) | Neal & Hulme, Lieberman, | ||
果皮Peel | DC (1R) + W (1T) | Hulme et al., | ||
果肉Pulp | DC (2R) | Romani et al., | ||
果肉Pulp | DC (2R) + DPGC + SPGC + W (2T) | Qin et al., | ||
梨 Pear | 果肉Pulp | DC (2R) | Romani et al., | |
DC (1R) + W (1T) + CSGC | Miller & Romani, | |||
DC (1R) + W (1T) | Li et al., | |||
核果类 Stone fruit | 鳄梨 Avocado | 果肉Pulp | DC (1R) + W (1T) | Biale et al., |
DC (3R) | Wiskich et al., | |||
DC (2R) | Lance et al., | |||
DC (1R) + W (1T) + SPGC + W (2T) | Moreau & Romani, | |||
芒果 Mango | 果肉Pulp | DC (2R) | Baqui et al., | |
DC (1R) + W (1T) | Kane & Marcellin, | |||
桃 Peach | 果肉Pulp | DC (1R) + W (1T) | Kan et al., | |
DC (1R) + W (2T) | Jin et al., | |||
DC (1R) + DPGC + W (1T) + SPGC + W (2T) | Qin et al., | |||
DC (1R) + DPGC + W (2T) | Yang et al., | |||
DC (1R) + W (1T) + DSGC + W (1T) | Jing et al., | |||
DC (1R) + DPGC + W (2T) | Wu et al., | |||
DC (1R) + DSGC + W (1T) | Wang et al., | |||
浆果类 Berries | 番茄 Tomato | 果肉Pulp | DC (1R) + W (1T) | Dickinson & Hanson, |
DC (2R) | Ku et al., | |||
DC (1R) + W (2T) | Li et al., | |||
DC (1R) + W (1T) + CSGC | Kalra & Brooks, | |||
DC (2R) + DPGC | Jeffery et al., | |||
DC (1R) + W (2T) + SPGC + W (2T) | Jarmuszkiewicz et al., | |||
DC (1R) + SPGC + W (2T) | Holtzapffel et al., | |||
DC (2R) + DPGC + W (2T) | de Oliveira et al., | |||
DC (1R) + DPGC + W (2T) + SPGC + W (2T) | Cai et al., | |||
香蕉 Banana | 果肉Pulp | DC (2R) | Haard & Hultin, | |
果皮Peel | DC (1R) + W (2T) + CSGC + W (1T) | Chotikakham et al., | ||
蓝莓 Blueberry | 果肉Pulp | DC (1R) + W (1T) | Zhou et al., | |
番木瓜Papaya 草莓Strawberry 番石榴Guava | 果肉Pulp | DC (2R) + DPGC + W (2T) | de Oliveira et al., | |
猕猴桃Kiwifruit | 果肉Pulp | DC (1R) + W (2T) | Ali et al., | |
菠萝 Pineapple | 果肉Pulp | DC (1R) + DSGC | Nukuntornprakit et al., | |
柑果类 Citrus | 柠檬Lemon | 果肉Pulp | DC (1R) + W (1T) | Bogin & Erickson, |
甜橙Orange | DC (2R) | Vines & Oberbacher, | ||
甜橙Orange | DC (1R) | Katz et al., | ||
柑Tangerine 橘Mandarin 橙Orange 柚Pummelo | DC (2R) + DPGC + W (2T) | Li et al., | ||
瓜类 Melon | 甜瓜 Muskmelon | 果肉Pulp | DC (2R) + DPGC + SPGC + W (2T) | Lyu et al., |
物质类别 Substrate categories | 代表物质及文献中报道的浓度(范围) Components and concentration | 作用及备注 Function and note |
---|---|---|
缓冲物质 Buffer material | 磷酸盐(Phosphate,0.01 ~ 0.50 mol · L-1),Tris(0.02 ~ 0.50 mol · L-1),Na-barbital(0.05 mol · L-1),Na4P2O7(0.025 mol · L-1),MOPS(0.05 ~ 0.50 mol · L-1),TES(0.025 mol · L-1),Bicine(0.1 mol · L-1),Hepes(0.01 ~ 0.04 mol · L-1) | 维持匀浆液pH稳定;根据果实样品特性调整缓冲物质的浓度;可同时使用多种缓冲物质 Keeping the pH of homogenate stable;Adjusting the concentration of buffer material according to the characteristics of the fruit samples;Multiple buffer materials can be used simultaneously |
渗透压调节物质 Osmolyte | 蔗糖(Sucrose,0.15 ~ 0.70 mol · L-1),甘露醇(Mannitol)/山梨醇(Sorbitol)(0.25 ~ 0.60 mol · L-1) | 调节溶液渗透压以维持线粒体膜结构稳定;可同时使用多种渗透压调节物质 Regulation of solution osmotic pressure to maintain the stability of mitochondrial membrane structure;Multiple osmolytes can be used simultaneously |
抗氧化物质 Antioxidant | 抗坏血酸(Ascorbic acid,0.01 ~ 0.03 mol · L-1),柠檬酸(Citric acid,0.02 mol · L-1),半胱氨酸(Cysteine,1 ~ 10 mmol · L-1或0.05% ~ 0.10% [w/v]),DTT(0.5 ~ 2.0 mmol · L-1),β-巯基乙醇(β-Mercaptoethanol,5 ~ 10 mmol L-1或0.1% [v/v]) | 维持多酚的还原性状态,延缓匀浆过程中的褐变;保护酶分子的还原性基团,稳定酶活性;用时现加 Maintaining the reducing state of polyphenols and delaying browning during homogenization;Protecting the reducing groups on the enzyme molecule and keeping the enzyme activity stable;Adding as needed |
螯合剂 Chelator | EDTA(0.2 ~ 10.0 mmol · L-1),EGTA(0.2 ~ 1.0 mmol · L-1) | 去除重金属离子对酶的抑制作用;浓度高时会抑制酶活性 Removing the inhibitory effect of heavy metal ions on enzymes;Inhibiting the enzyme activity at high concentrations |
其他 Other | PVP(0.05% ~ 5.00% [w/v]),PVPP(1.5% [w/v]) | 结合多酚以降低其对线粒体酶活性的抑制 Binding polyphenols to reduce the inhibition on mitochondrial enzyme activity |
BSA(0.05% ~ 1.00% [w/v]) | 降低脂肪酸对线粒体活性的抑制 Reducing the inhibition of fatty acids on mitochondrial enzyme activity | |
MgCl2(1 ~ 6 mmol · L-1),KCl(6 ~ 10 mmol · L-1),MgSO4(2 mmol · L-1) | 低浓度时可降低线粒体聚集且有利于酶活性,但高浓度时会加剧线粒体聚集 Reducing mitochondrial aggregation and promoting the enzyme activity at low concentrations;Increasing mitochondrial aggregation at high concentrations |
Table 2 The extraction buffer components for fruit mitochondria isolation
物质类别 Substrate categories | 代表物质及文献中报道的浓度(范围) Components and concentration | 作用及备注 Function and note |
---|---|---|
缓冲物质 Buffer material | 磷酸盐(Phosphate,0.01 ~ 0.50 mol · L-1),Tris(0.02 ~ 0.50 mol · L-1),Na-barbital(0.05 mol · L-1),Na4P2O7(0.025 mol · L-1),MOPS(0.05 ~ 0.50 mol · L-1),TES(0.025 mol · L-1),Bicine(0.1 mol · L-1),Hepes(0.01 ~ 0.04 mol · L-1) | 维持匀浆液pH稳定;根据果实样品特性调整缓冲物质的浓度;可同时使用多种缓冲物质 Keeping the pH of homogenate stable;Adjusting the concentration of buffer material according to the characteristics of the fruit samples;Multiple buffer materials can be used simultaneously |
渗透压调节物质 Osmolyte | 蔗糖(Sucrose,0.15 ~ 0.70 mol · L-1),甘露醇(Mannitol)/山梨醇(Sorbitol)(0.25 ~ 0.60 mol · L-1) | 调节溶液渗透压以维持线粒体膜结构稳定;可同时使用多种渗透压调节物质 Regulation of solution osmotic pressure to maintain the stability of mitochondrial membrane structure;Multiple osmolytes can be used simultaneously |
抗氧化物质 Antioxidant | 抗坏血酸(Ascorbic acid,0.01 ~ 0.03 mol · L-1),柠檬酸(Citric acid,0.02 mol · L-1),半胱氨酸(Cysteine,1 ~ 10 mmol · L-1或0.05% ~ 0.10% [w/v]),DTT(0.5 ~ 2.0 mmol · L-1),β-巯基乙醇(β-Mercaptoethanol,5 ~ 10 mmol L-1或0.1% [v/v]) | 维持多酚的还原性状态,延缓匀浆过程中的褐变;保护酶分子的还原性基团,稳定酶活性;用时现加 Maintaining the reducing state of polyphenols and delaying browning during homogenization;Protecting the reducing groups on the enzyme molecule and keeping the enzyme activity stable;Adding as needed |
螯合剂 Chelator | EDTA(0.2 ~ 10.0 mmol · L-1),EGTA(0.2 ~ 1.0 mmol · L-1) | 去除重金属离子对酶的抑制作用;浓度高时会抑制酶活性 Removing the inhibitory effect of heavy metal ions on enzymes;Inhibiting the enzyme activity at high concentrations |
其他 Other | PVP(0.05% ~ 5.00% [w/v]),PVPP(1.5% [w/v]) | 结合多酚以降低其对线粒体酶活性的抑制 Binding polyphenols to reduce the inhibition on mitochondrial enzyme activity |
BSA(0.05% ~ 1.00% [w/v]) | 降低脂肪酸对线粒体活性的抑制 Reducing the inhibition of fatty acids on mitochondrial enzyme activity | |
MgCl2(1 ~ 6 mmol · L-1),KCl(6 ~ 10 mmol · L-1),MgSO4(2 mmol · L-1) | 低浓度时可降低线粒体聚集且有利于酶活性,但高浓度时会加剧线粒体聚集 Reducing mitochondrial aggregation and promoting the enzyme activity at low concentrations;Increasing mitochondrial aggregation at high concentrations |
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