A gene expression reporter system (pHT3) for Clostridium acetobutylicum ATCC 824 was developed by using the lacZ gene from Thermoanaerobacterium thermosulfurogenes EM1 as the reporter gene. In order to test the reporter system, promoters of three key metabolic pathway genes, ptb (coding for phosphotransbutyrylase), thl (coding for thiolase), and adc (coding for acetoacetate decarboxylase), were...

We have used the bacterial beta-galactosidase gene (lacZ) as a reporter gene for the rapid measurement of T-cell antigen receptor (TCR)-mediated activation of individual T cells. The reporter construct contained the lacZ gene under the control of the nuclear factor of activated T cells (NF-AT) element of the human interleukin 2 enhancer [Fiering, S., Northrop, J. P., Nolan, G. P., Matilla, P., ...

BACKGROUND One of the main challenges for heterologous protein production by the methylotrophic yeast Pichia pastoris at large-scale is related to its high oxygen demand. A promising solution is a co-feeding strategy based on a methanol/sorbitol mixture during the induction phase. Nonetheless, a deep understanding of the cellular physiology and the regulation of the AOX1 promoter, used to gover...

Molecular dissection of mechanisms that govern the differentiated cardiac phenotype has, for cogent technical reasons, largely been undertaken to date in neonatal ventricular myocytes. To circumvent expected limitations of other methods, the present study was initiated to determine whether replication-deficient adenovirus would enable efficient gene transfer to adult cardiac cells in culture. A...

A clinical isolate of Mycobacterium avium was transformed with a new shuttle plasmid containing the Escherichia coli beta-galactosidase reporter gene under the control of the Mycobacterium bovis bacillus Calmette-Guérin (BCG) hsp60 promoter. beta-Galactosidase activity was assayed spectrophotometrically in bacterial homogenates of the recombinant strain (M. avium::lacZ) and used for quantificat...

Our study had three objectives: to extend the plasmid-based transformation protocol to a clinical isolate of C. trachomatis belonging to the trachoma biovar, to provide "proof of principle" that it is possible to "knock out" selected plasmid genes (retaining a replication competent plasmid) and to investigate the plasticity of the plasmid. A recently developed, plasmid-based transformation prot...

We report on a general strategy for engineering dominant negative mutations that, in principle, requires neither extensive structural or functional knowledge of the targeted protein. The approach consists of fusing the lysosomal protease cathepsin B (CB) to a subunit of a multimeric protein. The CB fusion polypeptide can proteolytically digest the multimer and/or detour the multimer from its us...

We have developed a model system for assessing the demethylating potential of external agents. Disruption in the DNA methylation pattern was evaluated at the translational level of the Escherichia coli beta-galactosidase coding gene (lacZ). We have constructed a clonal cell line (A4/4 cells) derived from the adenovirus-transformed human embryonic kidney 293 strain. The A4/4 cells contain the E....