ABSTRACT
A recombinant strain Escherichia coli DH5alpha(pMD19-glnA) including Bacillus subtilis glnA gene was constructed. Capillary electrophoresis and nuclear magnetic resonance were used to determine qualitatively the product of transformation by recombinant strain, and the relative level of mRNA expression of glnA was also determined by fluorescence quantitative RT-PCR. Subsequently, SDS-PAGE (polyacrylamide gel electrophoresis) was used to analysis the relative level of protein. Surprisingly, there was no increase of glutamine production in this recombinant strain, but an obvious increase in the GABA (gamma-aminobutyric acid ) production. It was showed in the experiment that protein expression of the glutamine synthetase did not increase, although glnA gene can be transcribed normally in this recombined strain. The phenomenon of exogenous glnA gene interfering metabolism of Escherichia coli was worthy of further study.
Subject(s)
Bacillus subtilis , Genetics , Bacterial Proteins , Genetics , Metabolism , Escherichia coli , Genetics , Metabolism , Glutamate-Ammonia Ligase , Genetics , Metabolism , RNA, Messenger , Genetics , Metabolism , Recombination, Genetic , gamma-Aminobutyric AcidABSTRACT
Aim To study the mechanism of andrographolide(AD) on the proliferation and apoptosis induction in human esophageal cancer Ec9706 cells.Methods The spectrometry was used to detect the activity of caspase-3 in human esophageal cancer Ec9706 cells treated with or without AD for 6 h,12 h and 18 h,and to detect the activity of caspase-8 and caspase-9 in human esophageal cancer Ec9706 cells treated with or without AD for 6 h.The influence of AD on the proliferation of Ec9706 cells after treatment with or without Z-VAD-FMK(a broad-spectrum caspase inhibitor) was determined by MTT method and the result was compared.The changes of gene expression levels of bcl-2 were determined by immunohistochemical method.Results The expression level of bcl-2 gene was obviously lower in the cells treated with AD(30 mg?L-1,P