Chemically inducible gene expression in seeds before testa rupture

2015 ◽  
Vol 25 (3) ◽  
pp. 345-352 ◽  
Author(s):  
Mariko Nonogaki ◽  
Taira Sekine ◽  
Hiroyuki Nonogaki
Keyword(s):  
Gene Expression ◽  
Function Analysis ◽  
Expression System ◽  
Basic Research ◽  
Inducible Expression ◽  
Upstream Sequence ◽  
Dependent Manner ◽  
Inducible Gene Expression ◽  
Inducible Expression System ◽  
Inducible Gene

AbstractImpermeability of the testa hinders efficient penetration of some small chemicals, such as transcriptional inhibitors, through the endosperm and the embryo during seed experiments. InArabidopsisseeds, 5-bromo-4-chloro-3-indolyl β-d-glucuronic acid, a substrate for β-glucuronidase, did not permeate through the endosperm and embryo efficiently at the stages before testa rupture. TheArabidopsistesta also limited efficient entry of methoxyfenozide, a chemical ligand that was used for inducible gene expression experiments, into seeds. While the detection of a reporter gene at the early imbibitional stages could be replaced by reverse transcription-polymerase chain reaction (RT-PCR), the interference of entry of the chemical ligand into seeds by the testa was still problematic to gene induction experiments. To develop an efficient inducible expression system for gene function analysis in seeds, an inducible expression system with nitrate, which is a testa-permeable ligand, was examined. The vector containing the 2.1-kb upstream sequence ofNITRITE REDUCTASE 1was able to cause expression of a test gene (long non-coding RNA) in imbibed seeds at the stage before testa rupture in a nitrate-dependent manner. This system can be used not only for characterization of genes associated with seed dormancy and germination in basic research, but also for the development of germination recovery or enhancement technologies for agricultural applications.

scholarly journals Tetracycline-Inducible Gene Expression and Gene Deletion in Candida albicans

2005 ◽  
Vol 4 (8) ◽  
pp. 1328-1342 ◽  
Author(s):  
Yang-Nim Park ◽  
Joachim Morschhäuser
Keyword(s):  
Gene Expression ◽  
Candida Albicans ◽  
Gene Function ◽  
High Efficiency ◽  
Expression System ◽  
Inducible Expression ◽  
Induced Expression ◽  
Inducible Gene Expression ◽  
Gene Expression System ◽  
Inducible Gene

ABSTRACT The genetic analysis of Candida albicans, the major fungal pathogen of humans, is hampered by its diploid genome, the absence of a normal sexual cycle, and a nonstandard codon usage. Although effective methods to study gene function have been developed in the past years, systems to control gene expression in C. albicans are limited. We have established a system that allows induction of gene expression in C. albicans by the addition of tetracycline (Tet). By fusing genetically modified versions of the reverse Tet repressor from Escherichia coli and the transcription activation domain of the Gal4 protein from Saccharomyces cerevisiae, a C. albicans-adapted reverse Tet-dependent transactivator (rtTA) was created that was expressed from the constitutive ADH1 or the opaque-specific OP4 promoter. To monitor Tet-inducible gene expression, the caGFP reporter gene was placed under the control of a Tet-dependent promoter, obtained by fusing a minimal promoter from C. albicans to seven copies of the Tet operator sequence. Fluorescence of the cells demonstrated that gene expression could be efficiently induced by the addition of doxycycline in yeast, hyphal, and opaque cells of C. albicans. The Tet-inducible gene expression system was then used to manipulate the behavior of the various growth forms of C. albicans. Tet-induced expression of a dominant-negative CDC42 allele resulted in growth arrest as large, multinucleate cells. Filamentous growth was efficiently inhibited under all tested hyphal-growth-promoting conditions by Tet-inducible expression of the NRG1 repressor. Tet-induced expression of the MTL a 1 gene in opaque cells of an MTLα strain forced the cells to switch to the white phase, whereas Tet-induced expression of the MTL a 2 transcription factor induced shmooing. When the ecaFLP gene, encoding the site-specific recombinase FLP, was placed under the control of the Tet-dependent promoter, Tet-inducible deletion of genes which were flanked by the FLP target sequences was achieved with high efficiency to generate conditional null mutants. In combination with the dominant selection marker caSAT1, the Tet-inducible gene expression system was also applied in C. albicans wild-type strains, including drug-resistant clinical isolates that overexpressed the MDR1, CDR1, and CDR2 multidrug efflux pumps. This system, therefore, allows a growth medium-independent, Tet-inducible expression and deletion of genes in C. albicans and provides a convenient, versatile new tool to study gene function and manipulate cellular behavior in this model pathogenic fungus.

scholarly journals Cumate-Inducible Gene Expression System for Sphingomonads and Other Alphaproteobacteria

2013 ◽  
Vol 79 (21) ◽  
pp. 6795-6802 ◽  
Author(s):  
Andreas Kaczmarczyk ◽  
Julia A. Vorholt ◽  
Anne Francez-Charlot
Keyword(s):  
Gene Expression ◽  
Caulobacter Crescentus ◽  
Paracoccus Denitrificans ◽  
Expression System ◽  
Model Organisms ◽  
Inducible Gene Expression ◽  
Gene Expression System ◽  
Content Type ◽  
Wide Range ◽  
Inducible Gene

ABSTRACTTunable promoters represent a pivotal genetic tool for a wide range of applications. Here we present such a system for sphingomonads, a phylogenetically diverse group of bacteria that have gained much interest for their potential in bioremediation and their use in industry and for which no dedicated inducible gene expression system has been described so far. A strong, constitutive synthetic promoter was first identified through a genetic screen and subsequently combined with the repressor and the operator sites of thePseudomonas putidaF1cym/cmtsystem. The resulting promoter, termed PQ5, responds rapidly to the inducer cumate and shows a maximal induction ratio of 2 to 3 orders of magnitude in the different sphingomonads tested. Moreover, it was also functional in otherAlphaproteobacteria, such as the model organismsCaulobacter crescentus,Paracoccus denitrificans, andMethylobacterium extorquens. In the noninduced state, expression from PQ5is low enough to allow gene depletion analysis, as demonstrated with the essential genephyPofSphingomonassp. strain Fr1. A set of PQ5-based plasmids has been constructed allowing fusions to affinity tags or fluorescent proteins.

scholarly journals Construction and Characterization of a Gradually Inducible Expression Vector for Halobacterium salinarum, Based on thekdpPromoter

2012 ◽  
Vol 78 (7) ◽  
pp. 2100-2105 ◽  
Author(s):  
Dorthe Kixmüller ◽  
Jörg-Christian Greie
Keyword(s):  
Gene Expression ◽  
Expression Vector ◽  
Expression System ◽  
Expression Patterns ◽  
Inducible Expression ◽  
Halobacterium Salinarum ◽  
Expression Vectors ◽  
Content Type ◽  
Inducible Expression System

ABSTRACTGradually inducible expression vectors which are governed by variations of growth conditions are powerful tools for gene expression of conditionally lethal mutants. Furthermore, controlled expression allows monitoring of overproduction of proteins at various stages in their expressing hosts. ForHalobacterium salinarum, which is often used as a paradigm for halophilic archaea, such an inducible expression system is not available to date. Here we show that thekdppromoter (Pkdp), which facilitates gene expression upon K+limitation, can be used to establish such a system for molecular applications. Pkdpfeatures a rather high expression rate, with an approximately 50-fold increase that can be easily varied by K+concentrations in the growth medium. Besides the construction of an expression vector, our work describes the characterization of expression patterns and, thus, offers a gradually inducible expression system to the scientific community.

scholarly journals Implementation of the agmatine-controlled expression system for inducible gene expression in Lactococcus lactis

2015 ◽  
Vol 14 (1) ◽  
Author(s):  
Daniel M. Linares ◽  
Patricia Alvarez-Sieiro ◽  
Beatriz del Rio ◽  
Victor Ladero ◽  
Begoña Redruello ◽  
...  
Keyword(s):  
Gene Expression ◽  
Lactococcus Lactis ◽  
Expression System ◽  
Inducible Gene Expression ◽  
Inducible Gene

Heat-Inducible Gene Expression System by Applying Alternating Magnetic Field to Magnetic Nanoparticles

2013 ◽  
Vol 3 (5) ◽  
pp. 273-279 ◽  
Author(s):  
Masaki Yamaguchi ◽  
Akira Ito ◽  
Akihiko Ono ◽  
Yoshinori Kawabe ◽  
Masamichi Kamihira
Keyword(s):  
Gene Expression ◽  
Magnetic Field ◽  
Magnetic Nanoparticles ◽  
Expression System ◽  
Alternating Magnetic Field ◽  
Inducible Gene Expression ◽  
Gene Expression System ◽  
Inducible Gene

scholarly journals A tunable anthranilate-inducible gene expression system for Pseudomonas species

2020 ◽  
Vol 105 (1) ◽  
pp. 247-258
Author(s):  
Lena Hoffmann ◽  
Michael-Frederick Sugue ◽  
Thomas Brüser
Keyword(s):  
Gene Expression ◽  
Expression System ◽  
Ant System ◽  
Plant Host ◽  
Inducible Gene Expression ◽  
Gene Expression System ◽  
Expression Levels ◽  
Key Points ◽  
Wide Range ◽  
Inducible Gene

Abstract Pseudomonads are among the most common bacteria in soils, limnic ecosystems, and human, animal, or plant host environments, including intensively studied species such as Pseudomonas aeruginosa, P. putida, or P. fluorescens. Various gene expression systems are established for some species, but there is still a need for a simple system that is suitable for a wide range of pseudomonads and that can be used for physiological applications, i.e., with a tuning capacity at lower expression levels. Here, we report the establishment of the anthranilate-dependent PantA promoter for tunable gene expression in pseudomonads. During studies on P. fluorescens, we constructed an anthranilate-inducible AntR/PantA-based expression system, named pUCP20-ANT, and used GFP as reporter to analyze gene expression. This system was compared with the rhamnose-inducible RhaSR/PrhaB-based expression system in an otherwise identical vector background. While the rhamnose-inducible system did not respond to lower inducer concentrations and always reached high levels over time when induced, expression levels of the pUCP20-ANT system could be adjusted to a range of distinct lower or higher levels by variation of anthranilate concentrations in the medium. Importantly, the anthranilate-inducible expression system worked also in strains of P. aeruginosa and P. putida and therefore will be most likely useful for physiological and biotechnological purposes in a wide range of pseudomonads. Key points • We established an anthranilate-inducible gene expression system for pseudomonads. • This system permits tuning of gene expression in a wide range of pseudomonads. • It will be very useful for physiological and biotechnological applications.

Development of an inflammation-inducible gene expression system using helper-dependent adenoviral vectors

2010 ◽  
Vol 12 (10) ◽  
pp. 832-839 ◽  
Author(s):  
Tianyao Yang ◽  
Rongqi Duan ◽  
Huibi Cao ◽  
Benjamin H. Lee ◽  
Chun Xia ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression System ◽  
Adenoviral Vectors ◽  
Inducible Gene Expression ◽  
Gene Expression System ◽  
Inducible Gene
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