Details
Original language | English |
---|---|
Pages (from-to) | 215-229 |
Number of pages | 15 |
Journal | Phyton - Annales Rei Botanicae |
Volume | 43 |
Issue number | 1 |
Publication status | Published - 21 Jul 2003 |
Abstract
It is a difficult task to isolate mitochondria from green tissue, especially from the widely used model plant Arabidopsis thaliana. However, for several physiological queries very pure and physiologically intact mitochondria are needed. From three plant species, Pisum sativum L., Nicotiana tabacum L. and Arabidopsis thaliana HEYNH., mitochondria were isolated according to different purification protocols described before or by an own optimised protocol. The methods involved differential centrifugation and density gradient centrifugation on self-generating gradients or step-gradients of Percoll. The quality of the purified mitochondria were tested by different methods to prove intactness of the membrane system, the physiological activity and the contamination by other organelles. In summary, it was possible to isolate mitochondria from the three different plant species according to all criteria investigated. However, the reliability of the organelle preparations and the purity and intactness varied between the methods. To demonstrate the quality of the purified mitochondria with respect to different traits a combination of several tests is recommendable. Determination of chlorophyll contents and enzyme activities, functional analysis of the respiratory chain, and the use of the powerful technique of Blue-native/Tricine SDS polyacrylamide gel electrophoresis are necessary to fully characterize the mitochondria. Especially combining a visual result by comparing the separation of protein complexes in gels with physiological measurements seems to be very useful for a comprehensive characterization of the organelle preparation.
Keywords
- Arabidopsis thaliana, Blue-native gel electrophoresis, Mitochondria, Nicotiana tabacum, Pisum sativum
ASJC Scopus subject areas
- Environmental Science(all)
- Global and Planetary Change
- Biochemistry, Genetics and Molecular Biology(all)
- Physiology
- Agricultural and Biological Sciences(all)
- Ecology, Evolution, Behavior and Systematics
- Agricultural and Biological Sciences(all)
- Plant Science
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In: Phyton - Annales Rei Botanicae, Vol. 43, No. 1, 21.07.2003, p. 215-229.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - How to Document the Purity of Mitochondria Prepared from Green Tissue of Pea, Tobacco and Arabidopsis thaliana
AU - Hausmann, Nicole
AU - Werhahn, Wolf
AU - Huchzermeyer, Bernhard
AU - Braun, Hans Peter
AU - Papenbrock, Jutta
PY - 2003/7/21
Y1 - 2003/7/21
N2 - It is a difficult task to isolate mitochondria from green tissue, especially from the widely used model plant Arabidopsis thaliana. However, for several physiological queries very pure and physiologically intact mitochondria are needed. From three plant species, Pisum sativum L., Nicotiana tabacum L. and Arabidopsis thaliana HEYNH., mitochondria were isolated according to different purification protocols described before or by an own optimised protocol. The methods involved differential centrifugation and density gradient centrifugation on self-generating gradients or step-gradients of Percoll. The quality of the purified mitochondria were tested by different methods to prove intactness of the membrane system, the physiological activity and the contamination by other organelles. In summary, it was possible to isolate mitochondria from the three different plant species according to all criteria investigated. However, the reliability of the organelle preparations and the purity and intactness varied between the methods. To demonstrate the quality of the purified mitochondria with respect to different traits a combination of several tests is recommendable. Determination of chlorophyll contents and enzyme activities, functional analysis of the respiratory chain, and the use of the powerful technique of Blue-native/Tricine SDS polyacrylamide gel electrophoresis are necessary to fully characterize the mitochondria. Especially combining a visual result by comparing the separation of protein complexes in gels with physiological measurements seems to be very useful for a comprehensive characterization of the organelle preparation.
AB - It is a difficult task to isolate mitochondria from green tissue, especially from the widely used model plant Arabidopsis thaliana. However, for several physiological queries very pure and physiologically intact mitochondria are needed. From three plant species, Pisum sativum L., Nicotiana tabacum L. and Arabidopsis thaliana HEYNH., mitochondria were isolated according to different purification protocols described before or by an own optimised protocol. The methods involved differential centrifugation and density gradient centrifugation on self-generating gradients or step-gradients of Percoll. The quality of the purified mitochondria were tested by different methods to prove intactness of the membrane system, the physiological activity and the contamination by other organelles. In summary, it was possible to isolate mitochondria from the three different plant species according to all criteria investigated. However, the reliability of the organelle preparations and the purity and intactness varied between the methods. To demonstrate the quality of the purified mitochondria with respect to different traits a combination of several tests is recommendable. Determination of chlorophyll contents and enzyme activities, functional analysis of the respiratory chain, and the use of the powerful technique of Blue-native/Tricine SDS polyacrylamide gel electrophoresis are necessary to fully characterize the mitochondria. Especially combining a visual result by comparing the separation of protein complexes in gels with physiological measurements seems to be very useful for a comprehensive characterization of the organelle preparation.
KW - Arabidopsis thaliana
KW - Blue-native gel electrophoresis
KW - Mitochondria
KW - Nicotiana tabacum
KW - Pisum sativum
UR - http://www.scopus.com/inward/record.url?scp=0042512002&partnerID=8YFLogxK
M3 - Article
AN - SCOPUS:0042512002
VL - 43
SP - 215
EP - 229
JO - Phyton - Annales Rei Botanicae
JF - Phyton - Annales Rei Botanicae
SN - 0079-2047
IS - 1
ER -