Research Mission and Profile
Molecular Signal Processing
Bioorganic Chemistry
Biochemistry of Plant Interactions
Cell and Metabolic Biology
Independent Junior Research Groups
Program Center MetaCom
Publications
Good Scientific Practice
Research Funding
Networks and Collaborative Projects
Symposia and Colloquia
Alumni Research Groups
Publications
Publications - Cell and Metabolic Biology
Publications
Genetic resources for the model plant Arabidopsis comprise mutant lines defective in almost any single gene in reference accession Columbia. However, gene redundancy and/or close linkage often render it extremely laborious or even impossible to isolate a desired line lacking a specific function or set of genes from segregating populations. Therefore, we here evaluated strategies and efficiencies for the inactivation of multiple genes by Cas9-based nucleases and multiplexing. In first attempts, we succeeded in isolating a mutant line carrying a 70 kb deletion, which occurred at a frequency of ~ 1.6% in the T2 generation, through PCR-based screening of numerous individuals. However, we failed to isolate a line lacking Lhcb1 genes, which are present in five copies organized at two loci in the Arabidopsis genome. To improve efficiency of our Cas9-based nuclease system, regulatory sequences controlling Cas9 expression levels and timing were systematically compared. Indeed, use of DD45 and RPS5a promoters improved efficiency of our genome editing system by approximately 25–30-fold in comparison to the previous ubiquitin promoter. Using an optimized genome editing system with RPS5a promoter-driven Cas9, putatively quintuple mutant lines lacking detectable amounts of Lhcb1 protein represented approximately 30% of T1 transformants. These results show how improved genome editing systems facilitate the isolation of complex mutant alleles, previously considered impossible to generate, at high frequency even in a single (T1) generation.
Publications
De-oiled rapeseed is a rich source of proteins and phenolic compounds. The phenolic compounds, namely sinapic acid derivatives (SAD), could occur as free sinapic acid, esterified (as sinapine, the choline ester of sinapic acid) and decarboxylated (as canolol) forms. Rapeseed protein preparations containing very low phenolic compounds have been the focus of our ongoing research. A precipitated rapeseed protein isolate is investigated for SAD such as sinapine, sinapoyl glucose, canolol using HPLC–DAD and LC–MS. Profile of the phenolic compounds of de-oiled rapeseed, press cakes and the precipitated protein isolate are compared. HPLC–DAD analysis indicated SAD; particularly sinapine is the main phenolic compound of all the substrates. The protein derivation process did not remarkably alter the profile of the investigated protein isolate.
This page was last modified on 27 Jan 2025 .
