Precise control of microbial gene expression resulting in a defined, fast, and homogeneous response is of utmost importance for synthetic bio(techno)logical applications. However, even broadly ...applied biotechnological workhorses, such as Corynebacterium glutamicum, for which induction of recombinant gene expression commonly relies on the addition of appropriate inducer molecules, perform moderately in this respect. Light offers an alternative to accurately control gene expression, as it allows for simple triggering in a noninvasive fashion with unprecedented spatiotemporal resolution. Thus, optogenetic switches are promising tools to improve the controllability of existing gene expression systems. In this regard, photocaged inducers, whose activities are initially inhibited by light-removable protection groups, represent one of the most valuable photoswitches for microbial gene expression. Here, we report on the evaluation of photocaged isopropyl-β-D-thiogalactopyranoside (IPTG) as a light-responsive control element for the frequently applied tac-based expression module in C. glutamicum. In contrast to conventional IPTG, the photocaged inducer mediates a tightly controlled, strong, and homogeneous expression response upon short exposure to UV-A light. To further demonstrate the unique potential of photocaged IPTG for the optimization of production processes in C. glutamicum, the optogenetic switch was finally used to improve biosynthesis of the growth-inhibiting sesquiterpene (+)-valencene, a flavoring agent and aroma compound precursor in food industry. The variation in light intensity as well as the time point of light induction proved crucial for efficient production of this toxic compound. IMPORTANCE: Optogenetic tools are light-responsive modules that allow for a simple triggering of cellular functions with unprecedented spatiotemporal resolution and in a noninvasive fashion. Specifically, light-controlled gene expression exhibits an enormous potential for various synthetic bio(techno)logical purposes. Before our study, poor inducibility, together with phenotypic heterogeneity, was reported for the IPTG-mediated induction of lac-based gene expression in Corynebacterium glutamicum. By applying photocaged IPTG as a synthetic inducer, however, these drawbacks could be almost completely abolished. Especially for increasing numbers of parallelized expression cultures, noninvasive and spatiotemporal light induction qualifies for a precise, homogeneous, and thus higher-order control to fully automatize or optimize future biotechnological applications.
Photocaged compounds are light-responsive biomolecules that regain their primal function upon short light exposure. Thus, cellular functions such as bacterial gene expression can be non-invasively ...triggered in a gradual and homogenous fashion by light. Especially for single-cell research, optogenetic tools exhibit an enormous potential for achieving an accurate and straightforward control of parallelized expression cultures as shown here with photocaged Isopropyl-β-
d
-thioalactopyranoside (IPTG) in
Escherichia coli
.
Light can be used to control numerous cellular processes including protein function and interaction as well as gene expression in a non-invasive fashion and with unprecedented spatiotemporal ...resolution. However, for chemical phototriggers tight, gradual, and homogeneous light response has never been attained in living cells. Here, we report on a light-responsive bacterial T7 RNA polymerase expression system based on a photocaged derivative of the inducer molecule isopropyl- beta -d-thiogalactopyranoside (IPTG). We have comparatively analyzed different Escherichia coli lacpromoter-regulated expression systems in batch and microfluidic single-cell cultivation. The lacY-deficient E. colistrain Tuner(DE3) harboring additional plasmid-born copies of the lacIgene exhibited a sensitive and defined response to increasing IPTG concentrations. Photocaged IPTG served as a synthetic photo-switch to convert the E. colisystem into an optogenetic expression module allowing for precise and gradual light-triggering of gene expression as demonstrated at the single cell level.
Provider: - Institution: - Data provided by Europeana Collections- Düsseldorf, Heinrich-Heine-Universität, Diss., 2012- All metadata published by Europeana are available free of restriction under the ...Creative Commons CC0 1.0 Universal Public Domain Dedication. However, Europeana requests that you actively acknowledge and give attribution to all metadata sources including Europeana
This article documents the addition of 83 microsatellite marker loci and 96 pairs of single‐nucleotide polymorphism (SNP) sequencing primers to the Molecular Ecology Resources Database. Loci were ...developed for the following species: Bembidion lampros, Inimicus japonicus, Lymnaea stagnalis, Panopea abbreviata, Pentadesma butyracea, Sycoscapter hirticola and Thanatephorus cucumeris (anamorph: Rhizoctonia solani). These loci were cross‐tested on the following species: Pentadesma grandifolia and Pentadesma reyndersii. This article also documents the addition of 96 sequencing primer pairs and 88 allele‐specific primers or probes for Plutella xylostella.
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