Universität zu Köln

Jaegle, Benjamin, Uroic, Miran Kalle, Holtkotte, Xu, Lucas, Christina, Termath, Andreas Ole, Schmalz, Hans-Guenther ORCID: 0000-0003-0489-1827, Bucher, Marcel ORCID: 0000-0003-1680-9413, Hoecker, Ute, Huelskamp, Martin and Schrader, Andrea ORCID: 0000-0002-3879-7057 (2016). A fast and simple LC-MS-based characterization of the flavonoid biosynthesis pathway for few seed(ling)s. BMC Plant Biol., 16. LONDON: BMC. ISSN 1471-2229

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Abstract

Background: (Pro)anthocyanidins are synthesized by the flavonoid biosynthesis pathway with multi-layered regulatory control. Methods for the analysis of the flavonoid composition in plants are well established for different purposes. However, they typically compromise either on speed or on depth of analysis. Results: In this work we combined and optimized different protocols to enable the analysis of the flavonoid biosynthesis pathway with as little as possible biological material. We chose core substances of this metabolic pathway that serve as a fingerprint to recognize alterations in the main branches of the pathway. We used a simplified sample preparation, two deuterated internal standards, a short and efficient LC separation, highly sensitive detection with tandem MS in multiple reaction monitoring (MRM) mode and hydrolytic release of the core substances to reduce complexity. The method was optimized for Arabidopsis thaliana seeds and seedlings. We demonstrate that one Col-0 seed/seedling is sufficient to obtain a fingerprint of the core substances of the flavonoid biosynthesis pathway. For comparative analysis of different genotypes, we suggest the use of 10 seed(lings). The analysis of Arabidopsis thaliana mutants affecting steps in the pathway revealed foreseen and unexpected alterations of the pathway. For example, HY5 was found to differentially regulate kaempferol in seeds vs. seedlings. Furthermore, our results suggest that COP1 is a master regulator of flavonoid biosynthesis in seedlings but not of flavonoid deposition in seeds. Conclusions: When sample numbers are high and the plant material is limited, this method effectively facilitates metabolic fingerprinting with one seed(ling), revealing shifts and differences in the pathway. Moreover the combination of extracted non-hydrolysed, extracted hydrolysed and non-extracted hydrolysed samples proved useful to deduce the class of derivative from which the individual flavonoids have been released.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Jaegle, Benjamin UNSPECIFIED UNSPECIFIED UNSPECIFIED Uroic, Miran Kalle UNSPECIFIED UNSPECIFIED UNSPECIFIED Holtkotte, Xu UNSPECIFIED UNSPECIFIED UNSPECIFIED Lucas, Christina UNSPECIFIED UNSPECIFIED UNSPECIFIED Termath, Andreas Ole UNSPECIFIED UNSPECIFIED UNSPECIFIED Schmalz, Hans-Guenther UNSPECIFIED orcid.org/0000-0003-0489-1827 UNSPECIFIED Bucher, Marcel UNSPECIFIED orcid.org/0000-0003-1680-9413 UNSPECIFIED Hoecker, Ute UNSPECIFIED UNSPECIFIED UNSPECIFIED Huelskamp, Martin UNSPECIFIED UNSPECIFIED UNSPECIFIED Schrader, Andrea UNSPECIFIED orcid.org/0000-0002-3879-7057 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-265387
DOI:	10.1186/s12870-016-0880-7
Journal or Publication Title:	BMC Plant Biol.
Volume:	16
Date:	2016
Publisher:	BMC
Place of Publication:	LONDON
ISSN:	1471-2229
Language:	English
Faculty:	Faculty of Mathematics and Natural Sciences
Divisions:	Faculty of Mathematics and Natural Sciences > Department of Biology > Botanical Institute
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language MYB TRANSCRIPTION FACTORS; FUNCTIONAL GENOMICS; GENE ENCODES; ANTHOCYANIN BIOSYNTHESIS; DOMAIN PROTEIN; ARABIDOPSIS; LIGHT; EXPRESSION; COP1; IDENTIFICATION Multiple languages Plant Sciences Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/26538