Format

Send to

Choose Destination
Biotechnol Bioeng. 2017 Sep;114(9):2011-2022. doi: 10.1002/bit.26321. Epub 2017 May 23.

Fast and reliable strain characterization of Streptomyces lividans through micro-scale cultivation.

Author information

1
Forschungszentrum Jülich GmbH, Institute of Bio- and Geosciences, IBG-1: Biotechnology, Leo-Brandt-Straße, 52428, Jülich, Germany.
2
Laboratory of Molecular Bacteriology, Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven-University of Leuven, Leuven, Belgium.
3
Center for Biotechnology (CeBiTec), Microbial Genomics and Biotechnology, Bielefeld University, Bielefeld, Germany.
4
Department of Molecular Biology Department, The National Research Centre, Dokki, Giza, Egypt.
5
Institute of Biotechnology, RWTH Aachen University, Worringer Weg 3, 52074, Aachen, Germany.

Abstract

Filamentous organisms of the genus Streptomyces play an important role in industrial production processes, due to their extensive secondary metabolism variability, as well as their ability to secrete efficiently large amounts of (heterologous) proteins. While genetic engineering tools are available to rapidly build up large strain libraries, the subsequent strain screening and bioprocess development still constitutes a bottleneck. This is due to the lack of reliable parallelized and accelerated cultivation techniques for morphologically challenging organisms. To address this challenge, we developed an integrated cultivation workflow for Streptomyces lividans based on a parallelized shaken 48-well microtiter-plate (MTP) cultivation device. In a first step, a feasible pre-culture method was identified and validated, revealing high comparability in subsequent main cultivations (coefficient of variation of 1.1% for in-plate replicates and 3.2% between different pre-cultures). When validating the growth performance in 1 mL MTP cultivation against an established 1,000 mL lab-scale cultivation system, highly comparable cultivation patterns were found for online (pH, dissolved oxygen), as well as for offline derived parameters (glucose uptake, cell-dry-weight, and pellet size). Additionally, the two cultivation regimes were compared with respect to transcriptional and protein secretion activity of Streptomyces, showing overall good comparability with minor, but well explainable discrepancies, most probably caused by different energy dissipation (shaking vs. stirring) and adaption effects due to different illumination conditions. Embedded within the presented cultivation workflow, the 1 mL MTP-based parallelized cultivation system seems to be a suitable screening tool for filamentous and industrial relevant organisms like Streptomyces. This can contribute to widen the field of application for these organisms and facilitate screening and early-stage bioprocess development. Biotechnol. Bioeng. 2017;114: 2011-2022.

KEYWORDS:

Streptomyces lividans TK24; filamentous morphology; high-throughput bioprocess development; miniaturized cultivation; parallelized cultivation; proteomics; strain characterization; transcriptomics

PMID:
28436005
DOI:
10.1002/bit.26321
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Wiley
Loading ...
Support Center