Based on the transcriptome analysis data of a Bacillus licheniformis, a novel bidirectional promoter was identified from the strain and its transcriptional strength was analyzed. The expression level of a Bacillus clausii derived alkaline protease gene driven by the bidirectional promoter was studied by using the known strong constitutive promoter pShuttle-09 as a control. Three recombinant expression vectors and the corresponding recombinant bacteria were constructed. Under the control of the new promoter pLA and its reverse promoter pLB, the alkaline protease expression level respectively reached 164 U/mL and 111 U/mL. The results indicated that the transcription strength of pLA was significantly higher than that of pShuttle-09 and pLB, and both the pLA and pLB promoters could initiate the expression of the alkaline protease. Thus, it provides a new expression element for the heterogenous genes in Bacillus sp. and a new idea for the co-expression of two genes in one prokaryotic strain.
Bacillus subtilis ; Promoter Regions, Genetic

Recently, the Encyclopedia of DNA Elements (ENCODE) Analysis Working Group converted data from ChIP-seq analyses from the Broad Histone track into 15 corresponding chromatic maps that label sequences with different kinds of histone modifications in promoter regions. Here, we publish a frequency profile of the three ChromHMM promoter states, at 200-bp intervals, with particular reference to the existence of sequence patterns of promoter elements, GC-richness, and transcription starting sites. Through detailed and diligent analysis of promoter regions, researchers will be able to uncover new and significant information about transcription initiation and gene function.
DNA ; Epigenomics ; Histones ; Promoter Regions, Genetic

No abstract available.
DNA* ; Genome* ; Nucleopolyhedrovirus* ; Promoter Regions, Genetic*

OBJECTIVETo observe the therapeutic effect of CDglyTK gene mediated by synthetic radiation-inducible promoter in the treatment of Tca8113 cells.

METHODSCDglyTK gene in pCEA-CDglyTK was subcloned into pcDNA3.1 (+) to construct plasmid pcDNA3.1 (+)-CDglyTK, and then the synthetic radiation-inducible promoter in pMD18 -T -E was inserted into pcDNA3.1 (+) -CDglyTK to construct plasmid pcDNA3.1 (+ )/E -CDglyTK. The recombinant plasmid was transfected into Tca8113 cells by lipofectamine, and then exposed to 3 Gy irradiation. Cytotoxicity was evaluated by MTT. The expression of CDglyTK gene was detected by RT-PCR. The apoptosis and proliferation were examined by flow cytomtery.

RESULTSThe plasmid pcDNA3.1 (+)/E-CDglyTK was constructed successfully. The comparative survival rate of Tca8113 cells was markedly decreased by induction irradiation. Up-regulation of CDglyTK expression was found in Tca8113 cells exposed to irradiation. The apoptosis index (AI) of Tca8113 cells exposed to irradiation was higher than that of Tca8113 cells without irradiation, the other way round, the proliferation index (PI) of Tca8113 cells exposed to irradiation was lower than that of Tca8113 cells without irradiation.

CONCLUSIONThe synthetic radiation-inducible promoter can be served as a molecular switch to improve the expression of CDglyTK gene in Tca8113 cells, and low dose induction radiation can significantly improve the therapeutic efficiency.

OBJECTIVE: Recently, many experimental evidences have been reported that psychiatric diseases are closely related with neurodevelopmental abnormalities and this can be properly explained by apoptosis. It is known that Apo-1/Fas is one of the genes in charge of apoptosis related with neurodevelopmental abnormalities. In this study, the association between bipolar disorder and functional polymorphism in Apo-1/Fas promoter gene has been investigated. METHOD: For 81 bipolar disorder patients and 217 healthy control subjects, MvaI restriction fragment length polymorphism(RFLP) of Apo-1/Fas promoter gene was analyzed after polymerase chain reaction(PCR) amplification. RESULT: There was a statistical significant difference in genotypic distribution(chi2=16.656, df=2, p=0.0002) and allelic frequencies(chi2=14.225, df=1, p=0.0002) between bipolar disorder patients and healthy control subjects. CONCLUSION: Our results suggest an association between functional polymorphism in Apo-1/Fas promoter gene and bipolar disorder and provide the important genetic information related with the pathogenesis of the disease. Further studies employing larger samples are required to clarify the present results.
Apoptosis ; Bipolar Disorder* ; Humans ; Promoter Regions, Genetic*

An efficient genetic transformation system and suitable promoters are essential prerequisites for gene expression studies and genetic engineering in streptomycetes. In this study, firstly, a genetic transformation system based on intergeneric conjugation was developed in Streptomyces rimosus M527, a bacterial strain which exhibits strong antagonistic activity against a broad range of plant-pathogenic fungi. Some experimental parameters involved in this procedure were optimized, including the conjugative media, ratio of donor to recipient, heat shock temperature, and incubation time of mixed culture. Under the optimal conditions, a maximal conjugation frequency of 3.05×10^-5 per recipient was obtained. Subsequently, based on the above developed and optimized transformation system, the synthetic promoters SPL-21 and SPL-57, a native promoter potrB, and a constitutive promoter permE^* commonly used for gene expression in streptomycetes were selected and their activity was analyzed using gusA as a reporter gene in S. rimosus M527. Among the four tested promoters, SPL-21 exhibited the strongest expression activity and gave rise to a 2.2-fold increase in β-glucuronidase (GUS) activity compared with the control promoter permE^*. Promoter SPL-57 showed activity comparable to that of permE^*. Promoter potrB, which showed the lowest activity, showed a 50% decrease in GUS activity compared with the control permE^*. The transformation system developed in this study and the tested promotors provide a basis for the further modification of S. rimosus M527.
Conjugation, Genetic ; Glucuronidase/genetics* ; Promoter Regions, Genetic ; Streptomyces rimosus/genetics*

DNA methylation and demethylation are two important mechanisms of epigenetics, which is important in the study of cell differentiation, proliferation, and senescence. During osteogenic differentiation of stem cells, the expression of osteogenic specific genes and demethylated promoters is upregulated, whereas the expression of pluripotent genes and hypermethylated promoters is downregulated. The dynamic changes and balance between DNA methylation and demethylation are important for the coordination of gene expression and the inhibition of improper phenotypes. Abnormal changes in the methylation modification mechanism in osteogenic differentiation not only affect the normal function of stem cells but are also associated with the occurrence and development of many common skeletal diseases. This paper reviews the new progress of DNA methylation and demethylation in regulating osteogenic differentiation. The possible skeletal diseases caused by abnormal DNA methylation are also presented.
Cell Differentiation ; DNA Methylation ; Osteogenesis ; Promoter Regions, Genetic ; Stem Cells

Apoptosis has been hypothesized to be involved in the pathogenesis in schizophrenia. A large number of genes are known to mediate the apoptotic process; Apo-1/Fas (CD95) is a well-known example of such genes. In the present study, MvaI restriction fragment length polymorphism, a polymorphic marker present within the Apo-1/Fas gene, was examined in a population consisting of 226 control subjects and 110 schizophrenia patients, all of them Korean in ethnicity. No statistically significant difference in the genotypic distribution and allelic frequencies was observed between the control and the schizophrenia patient group. To find out the precise effect of Apo-1/Fas gene polymorphisms on the susceptibility to schizophrenia, further studies are warranted to investigate possible involvement of other polymorphisms with a larger sample population.
Apoptosis ; Humans ; Polymorphism, Restriction Fragment Length ; Promoter Regions, Genetic ; Schizophrenia*

We constructed bicistronic expression system containing AH6 promoter, 5' UTR and its fore 38 bp sequence from Corynebacterium glutamicum, followed by a conserved Shine-Dalgarno (SD) sequence for xylanase expression. The two major secretory pathways signal peptide in C. glutamicum, Tat (CgR0949) and Sec (CspB) dependent signal peptide were added before xylanase for its secretion. Fed-batch cultivation was done in a 5 L jar for high-level xylanase secretion. The enzyme properties of the purified xylanase were then studied, including the effect of temperature and pH on its activity. The xylanase could be secreted into the culture supernatant when the Sec-dependent signal peptide CspB was used, but none was detected when CgR0949 was used. The secretory production level of xylanase in a flask was 486.2 U/mL and become 1 648.7 U/mL when in a 5 L jar, which was 3.4 fold as in the flask. The optimal pH and temperature of xylanase were pH 4.5 and 45 ℃, respectively. Its activity was 80% of initial activity after pretreatment at 4 ℃ for 24 h at pH 4-11, 95% after incubation below 50 ℃ for 15 min, and 20% when the temperature above 60 ℃. The xylanase could be efficiently secreted into the culture medium by C. glutamicum using its own genetic elements, and the secretion level could be improved through large-scale fed-batch cultivation. This bicistronic expression system can provide a useful tool for heterologous proteins secretion in C. glutamicum. In addition, the catalyze activity of xylanase could be further improved by enzyme properties study.
Corynebacterium glutamicum ; Promoter Regions, Genetic ; Protein Sorting Signals ; Protein Transport

Bacillus subtilis can be widely used as an important microorganism for metabolic engineering and recombinant proteins expression in industrial biotechnology and synthetic biology. However, it is difficult to make accurate regulation of exogenous gene by biological tools in B. subtilis, which limits the application of B. subtilis in synthetic biology. The purpose of this study is to develop regulatory tools for precise control of gene expression by using non-coding RNAs, by which the activation of heterologous gene could be achieved without the auxiliary protein factors. We constructed the synthetic riboswitch E and aptazyme AZ using the theophylline aptamer. Six different native promoters from B. subtilis were functionally adapted with the E and AZ to fabricate an array of novel regulatory elements activated by theophylline. Then, we determined the performance of these elements using green fluorescence protein as reporter, and then further verified using red fluorescence protein and pullulanase as cargo proteins. Results showed that the same kind of RNA elements with different promoters showed different levels of efficiency. Promoter PsigW and E combination (sigWE) had the highest induction rate in B. subtilis. Compared with the control group, it can produce the induction rate of 16.8. Promoter PrpoB and AZ combination (rpoBAZ) showed the highest induction rate of 6.2. SigWE mediated mCherry induction rate was 9.2, and P43E mediated pullulanase induction rate was 32.8, in which enzyme activity reached 81 U/mL. This study confirmed that GFP, mCherry and pullulan can all be regulated by riboswitch and aptazyme, but there were differences between different combinations of promoters with RNA regulators.
Bacillus subtilis ; Promoter Regions, Genetic ; RNA ; Recombinant Proteins ; Theophylline