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Chassis organism from Corynebacterium glutamicum – a top‐down approach to identify and delete irrelevant gene clusters
Author(s) -
Unthan Simon,
Baumgart Meike,
Radek Andreas,
Herbst Marius,
Siebert Daniel,
Brühl Natalie,
Bartsch Anna,
Bott Michael,
Wiechert Wolfgang,
Marin Kay,
Hans Stephan,
Krämer Reinhard,
Seibold Gerd,
Frunzke Julia,
Kalinowski Jörn,
Rückert Christian,
Wendisch Volker F.,
Noack Stephan
Publication year - 2015
Publication title -
biotechnology journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.144
H-Index - 84
eISSN - 1860-7314
pISSN - 1860-6768
DOI - 10.1002/biot.201400041
Subject(s) - corynebacterium glutamicum , biology , gene , prophage , chassis , genome , organism , computational biology , gene cluster , gene knockout , genetics , model organism , strain (injury) , bacteriophage , escherichia coli , structural engineering , engineering , anatomy
For synthetic biology applications, a robust structural basis is required, which can be constructed either from scratch or in a top‐down approach starting from any existing organism. In this study, we initiated the top‐down construction of a chassis organism from Corynebacterium glutamicum ATCC 13032, aiming for the relevant gene set to maintain its fast growth on defined medium. We evaluated each native gene for its essentiality considering expression levels, phylogenetic conservation, and knockout data. Based on this classification, we determined 41 gene clusters ranging from 3.7 to 49.7 kbp as target sites for deletion. 36 deletions were successful and 10 genome‐reduced strains showed impaired growth rates, indicating that genes were hit, which are relevant to maintain biological fitness at wild‐type level. In contrast, 26 deleted clusters were found to include exclusively irrelevant genes for growth on defined medium. A combinatory deletion of all irrelevant gene clusters would, in a prophage‐free strain, decrease the size of the native genome by about 722 kbp (22%) to 2561 kbp. Finally, five combinatory deletions of irrelevant gene clusters were investigated. The study introduces the novel concept of relevant genes and demonstrates general strategies to construct a chassis suitable for biotechnological application.