Purification of pea nodule symbiosomes using an aqueous polymer two-phase system.
Symbiosomes were obtained from mature pea (Pisum sativum cv. Argona) root nodules infected with Rhizobium leguminosarum strain (biov. viciae 3841) and purified using an aqueous polymer two-phase system (APS). The APS consists of a mixture of polymers, usually dextran T500 and poly(ethylene glycol) 3350, prepared as aqueous solutions on a weight per weight basis, where each fraction distributes according to their surface characteristics. Results of ATPase activity, cytochrome c oxidase activity, glucan synthase II activity, NAD(P)H-cytochrome c reductase activity, NO3(-)-sensitive ATPase activity, transport of [14C]malate vs. [14C]glutamate and MAC 57 antigen analysis showed that the APS method provided intact symbiosomes with low bacteroid, plasma membrane, endoplasmic reticulum and/or mitochondria contamination. No complicated equipment is needed and the method was simple and fast, compared with other purification techniques.
Hernández
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LE
Departamento de Química Agrícola, Geología y Geoquímica, Facultad de Ciencias, Universidad Autónoma de Madrid, Spain.
Rojas-Ojeda
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Cooke
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Carpena-Ruiz
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D000251:Adenosine Triphosphatases / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry / Q000382:microbiology / Q000648:ultrastructure; D002499:Centrifugation, Density Gradient; D003911:Dextrans / Q000737:chemistry; D003576:Electron Transport Complex IV / Q000378:metabolism; D015151:Immunoblotting; D018532:Pisum sativum / Q000737:chemistry / Q000382:microbiology* / Q000648:ultrastructure; D018517:Plant Roots / Q000737:chemistry / Q000382:microbiology* / Q000648:ultrastructure; D011092:Polyethylene Glycols / Q000737:chemistry; D011108:Polymers / Q000737:chemistry; D016961:Rhizobium leguminosarum / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D012997:Solvents / Q000737:chemistry; D013499:Surface Properties; D013559:Symbiosis
D003911:Dextrans; D011108:Polymers; D012997:Solvents; D011092:Polyethylene Glycols; D003576:Electron Transport Complex IV; D000251:Adenosine Triphosphatases