High-throughput method for Oxygen Consumption Rate measurement (OCR) in plant mitochondria
Abstract Background Conventional methods to measure oxygen consumption, such as Clark-type electrodes, have limitations such as requiring a large amount of starting material. Moreover, commercially available kits for high-throughput methods are usually optimized for animal cells and mitochondria. He...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
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BMC
2023-10-01
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| Series: | BMC Plant Biology |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s12870-023-04516-0 |
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| author | Hanna Fuchs Arleta Malecka Adrianna Budzinska Wieslawa Jarmuszkiewicz Liliana Ciszewska Aleksandra M. Staszak Joanna Kijowska-Oberc Ewelina Ratajczak |
| author_facet | Hanna Fuchs Arleta Malecka Adrianna Budzinska Wieslawa Jarmuszkiewicz Liliana Ciszewska Aleksandra M. Staszak Joanna Kijowska-Oberc Ewelina Ratajczak |
| author_sort | Hanna Fuchs |
| collection | DOAJ |
| description | Abstract Background Conventional methods to measure oxygen consumption, such as Clark-type electrodes, have limitations such as requiring a large amount of starting material. Moreover, commercially available kits for high-throughput methods are usually optimized for animal cells and mitochondria. Here, we present a novel method to measure the oxygen consumption rate using a high-throughput assay in isolated mitochondria of European beech seeds. To perform the measurements, we adapted the Agilent Seahorse XF Cell Mito Stress Test Kit protocol for measurements on plant mitochondria. Results The optimized protocol for OCR measurement of mitochondria isolated from beech seeds allowed the observation of storage period-dependent gradual decreases in non-phosphorylating respiration, phosphorylating respiration and maximal FCCP-stimulated respiration. The longer the seeds were stored, the greater the impairment of respiratory function. Conclusions Thanks to this method it is possible to minimize the amount of plant material and conduct research to obtain information on the respiratory condition and activity of plant mitochondria, including the efficiency of oxidative phosphorylation and the maximum oxidative capacity of the respiratory chain. We demonstrated that the improved protocol is suitable for study of plant material. |
| first_indexed | 2024-03-10T22:03:53Z |
| format | Article |
| id | doaj.art-ff8eaa0fde274a84a024148a20733a30 |
| institution | Directory Open Access Journal |
| issn | 1471-2229 |
| language | English |
| last_indexed | 2024-03-10T22:03:53Z |
| publishDate | 2023-10-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Plant Biology |
| spelling | doaj.art-ff8eaa0fde274a84a024148a20733a302023-11-19T12:52:58ZengBMCBMC Plant Biology1471-22292023-10-012311510.1186/s12870-023-04516-0High-throughput method for Oxygen Consumption Rate measurement (OCR) in plant mitochondriaHanna Fuchs0Arleta Malecka1Adrianna Budzinska2Wieslawa Jarmuszkiewicz3Liliana Ciszewska4Aleksandra M. Staszak5Joanna Kijowska-Oberc6Ewelina Ratajczak7Institute of Dendrology, Polish Academy of SciencesDepartment of Epidemiology and Cancer Prevention, Greater Poland Cancer CentreDepartment of Bioenergetics, Faculty of Biology, Adam Mickiewicz UniversityDepartment of Bioenergetics, Faculty of Biology, Adam Mickiewicz UniversityInstitute of Molecular Biology and Biotechnology, Faculty of Biology, Adam Mickiewicz University in PoznanLaboratory of Plant Physiology, Department of Plant Biology and Ecology Faculty of Biology, University of BialystokInstitute of Dendrology, Polish Academy of SciencesInstitute of Dendrology, Polish Academy of SciencesAbstract Background Conventional methods to measure oxygen consumption, such as Clark-type electrodes, have limitations such as requiring a large amount of starting material. Moreover, commercially available kits for high-throughput methods are usually optimized for animal cells and mitochondria. Here, we present a novel method to measure the oxygen consumption rate using a high-throughput assay in isolated mitochondria of European beech seeds. To perform the measurements, we adapted the Agilent Seahorse XF Cell Mito Stress Test Kit protocol for measurements on plant mitochondria. Results The optimized protocol for OCR measurement of mitochondria isolated from beech seeds allowed the observation of storage period-dependent gradual decreases in non-phosphorylating respiration, phosphorylating respiration and maximal FCCP-stimulated respiration. The longer the seeds were stored, the greater the impairment of respiratory function. Conclusions Thanks to this method it is possible to minimize the amount of plant material and conduct research to obtain information on the respiratory condition and activity of plant mitochondria, including the efficiency of oxidative phosphorylation and the maximum oxidative capacity of the respiratory chain. We demonstrated that the improved protocol is suitable for study of plant material.https://doi.org/10.1186/s12870-023-04516-0Oxygen consumption rateOCRHigh-throughput plant mitochondria respiration assayPlant mitochondria |
| spellingShingle | Hanna Fuchs Arleta Malecka Adrianna Budzinska Wieslawa Jarmuszkiewicz Liliana Ciszewska Aleksandra M. Staszak Joanna Kijowska-Oberc Ewelina Ratajczak High-throughput method for Oxygen Consumption Rate measurement (OCR) in plant mitochondria BMC Plant Biology Oxygen consumption rate OCR High-throughput plant mitochondria respiration assay Plant mitochondria |
| title | High-throughput method for Oxygen Consumption Rate measurement (OCR) in plant mitochondria |
| title_full | High-throughput method for Oxygen Consumption Rate measurement (OCR) in plant mitochondria |
| title_fullStr | High-throughput method for Oxygen Consumption Rate measurement (OCR) in plant mitochondria |
| title_full_unstemmed | High-throughput method for Oxygen Consumption Rate measurement (OCR) in plant mitochondria |
| title_short | High-throughput method for Oxygen Consumption Rate measurement (OCR) in plant mitochondria |
| title_sort | high throughput method for oxygen consumption rate measurement ocr in plant mitochondria |
| topic | Oxygen consumption rate OCR High-throughput plant mitochondria respiration assay Plant mitochondria |
| url | https://doi.org/10.1186/s12870-023-04516-0 |
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