Gene expression exhibits temporal and spatial patterns to response environmental changes and growth cycle. Gene expression is under strict control at different levels among which control at transcription level is the predominant mode, especially in prokaryotes. In this review, we summarized the new developments of methods used in transcriptional studies, including modifications and improvements of the classic methods, such as gel-shift assay, DNA foot printing, and in vivo reporter system. In addition, we introduced examples to apply new methods, such as surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC) to characterize protein-DNA, ligand-protein, and ligand-protein-DNA interactions. The collection of these methods and their application could guide and accelerate relevant studies.
Calorimetry ; DNA Footprinting ; Gene Expression ; Ligands ; Proteins ; Surface Plasmon Resonance ; Transcription, Genetic

Bacterial two-hybrid system is a newly developed method for studying protein-protein interactions. However, in our studies of the interaction of regulatory proteins in Streptomyces, it was found that the bacterial two-hybrid system is not sensitive enough by the blue-and-white selection on X-gal plate. To overcome this drawback, the reason of false positive clone was firstly determined, which was the disturbance of other direct or indirect regulation on lacZ promoter. Then the disturbance was diluted by introducing multicopy lacZ promoter, which drive another reporter gene gfp. By such design, the sensitivity of the modified bacterial two-hybrid system was significantly inproved and the two different reporters also help to decrease the rate of the false positive clones. Further the evaluation of the modifiedd bacterial two-hybrid system indicated that the sensitivity was significantly improved.
Bacteria ; Genes, Reporter ; Promoter Regions, Genetic ; Protein Interaction Mapping ; methods ; Two-Hybrid System Techniques

Objective To investigate the distribution characteristics of pathogenic bacteria among different seasons and ages in children lower respiratory tract infection.Methods To retrospectively analyze the results of bacterial culture in pediatric outpatients and inpatients with lower respiratory tract infections in our hospital from September 2014 to August 2015.The pathogenic bacterial detection situation of lower respiratory tract infection among different age groups and different were statistically analyzed.Results A total of 2 809 pathogenic bacterial strains were isolated from 4 629 lower respiratory tract specimens,and detection rate was 60.7%.The pathogenic bacteria distribution was different among different age groups.Haemophilus influenzae (33.3%) ranked the first place in the lower respiratory infection among children aged from 19 d to 7 year old.The detection rates of Haemophilus parainfluenzae,Moraxella catarrhalis and staphylococcus aureus had obvious seasonality,while Streptococcus pneumoniae had no obvious seasonality.Conclusion Pathogenic bacteria causing lower respiratory tract infections in children are different from those in adults,which are affected by age and seasonal change.

AIM: To investigate the synergistic induction of apoptosis in rhabdomyosarcoma cells by the combination of TRAIL or TRAIL gene with cisplatin. METHODS: Rhabdomyosarcoma cells were treated with TRAIL, (Ad/GT)-TRAIL, cisplatin, respectively or the combination for 3 days. The cytotoxicity was observed by MTT assay. The apoptotic rates and the expression rates of Fas protein were measured by flow cytometry (FCM). The expression of cFLIP mRNA was determined by RT-PCR. RESULTS: Rhabdomyosarcoma cells were treated with Ad/ GT-TRAIL and TRAIL (100.0 ?g/L), the cytotoxicity index were 52.5% and 43.5%, the percentage of apoptotic cells were 12.95% and 10.26%, respectively. Combined with cisplatin, the cytotoxicity index and the percentage of apoptotic cells were increased significantly (P

BACKGROUND:Clinical studies have demonstrated that systemic mild hypothermia could significantly reduce the disability and mortality rate of patients with severe traumatic brain injury. In recent years, the researches about the treatment of spinal cord injury by mild hypothermia have been successively carried out. OBJECTIVE:To investigate the effects of mild hypothermia on nerve regeneration microenvironment after spinal cord injury and explore the possible underlying mechanism of nerve regeneration and functional recovery after spinal cord injury in rats. METHODS:Twenty out of sixty-seven rats were randomly selected as the sham group, and other rats were used to establish spinal cord injury models in T9 segment using modified Alen's method. Three rats were excluded for failure in spinal cord injury induction and four rats for death during modeling. The rest 40 rats were randomly and evenly divided into the spinal cord injury and mild hypothermia groups (n=20 rats/group). The rats in the sham and spinal cord injury groups were placed in the operating table with normal temperature, making their rectal temperature at 37.0±0.5℃ for 72 hours. The rats in the mild hypothermia group were placed on ice blanket machine, making their rectal temperature at 34.0±0.5℃ for 72 hours, then the temperature was naturaly rewarmed. RESULTS AND CONCLUSION:Compared with spinal cord injury group, the apoptosis index and the level of aquaporin 4/9 mRNA and protein expression in spinal cord injury tissue were al decreased, somatosensory evoked potential latency and amplitude were recovered, and the motor functional scores were increased in the mild hypothermia group. These results indicate that mild hypothermia play its protective effect on spinal cord injury through attenuating apoptosis of neural cels and decreasing aquaporin 4/9 mRNA and protein expression levels.

Objective To investigate the antibiotic resistance of Haemophilus influenzae isolates collected from the children with respiratory tract infection for rational use of antibiotics in clinical practice .Methods The H .influenzae strains were isolated from children and subjected to antimicrobial susceptibility testing by Kirby-Bauer method .Nitrocefin disc test was used to detect the production of beta-lactamases .WHONET 5 .6 software was used to analyze the susceptibility data .Results A totalof1256strainsof H.influenzaewereisolated.About37.8% ,65.5% and16.5% ofthe1256strainsof H.influenzae were resistant to ampicillin ,trimethoprim-sulfamethoxazole ,and ampicillin-sulbactam ,respectively .Less than 10 .0% of these strains were resistant to any other antibiotics tested .Beta-lactamase was produced in 33 .5% of the 1 256 strains of H . influenzae .Conclusions The H . influenzae strains in this study are mainly resistant ampicillin and trimethoprim-sulfamethoxazole .About 80 .0% of these H . influenzae strains were still susceptible to cefaclor ,ampicillin-sulbactam , cefixime ,ceftazidime ,azithromycin ,ciprofloxacin ,meropenem and rifampin .The primary mechanism of ampicillin resistance in Haemophilus is production of beta-lactamases .

Objective To understand the status quo of Salmonella infection in the patients with infectious diarrhea in Zhongshan city .Methods The fecal samples collected from 6 417 outpatients or inpatients with infection diarrhea in our hospital from January 2010 to December 2013 were performed the Salmonella culture ,isolation and identification and at the same time the serological clas-sification and the pulsed-field gel electrophoresis (PFGE) were performed .Results Among 6 417 samples ,460 strains of Salmonel-la were detected with the isolation rate of 7 .17% .Salmonella infection was dominated by children infection .The Salmonella infec-tion rate in children aged less than 1 years was 52 .4% (241 cases) ,followed by children aged 1- < 3 years ,which was 36 .7% (169 cases) .In the serological classification ,the most of detected Salmonella was salmonella typhimurium 46 .1% ,accounting for 46 .1% , Stanley salmonella accounting for 13 .5% and Salmonella enteritidis accounting for 8 .7% .The PFGE homology analysis showed that in the XbaI enzyme digestion spectral band ,26 PFGE band types had 100% homology .Conclusion Salmonella is one of the main pathogenic bacteria causing diarrhea and serious impact on health ,which is worthy of clinicians to pay attention to .

Streptomycetes produce many antibiotics and are important model microorgansims for scientific research and antibiotic production. Metabolomics is an emerging technological platform to analyze low molecular weight metabolites in a given organism qualitatively and quantitatively. Compared to other Omics platform, metabolomics has greater advantage in monitoring metabolic flux distribution and thus identifying key metabolites related to target metabolic pathway. The present work aims at establishing a rapid, accurate sample preparation protocol for metabolomics analysis in streptomycetes. In the present work, several sample preparation steps, including cell quenching time, cell separation method, conditions for metabolite extraction and metabolite derivatization were optimized. Then, the metabolic profiles of Streptomyces coelicolor during different growth stages were analyzed by GC-MS. The optimal sample preparation conditions were as follows: time of low-temperature quenching 4 min, cell separation by fast filtration, time of freeze-thaw 45 s/3 min and the conditions of metabolite derivatization at 40 degrees C for 90 min. By using this optimized protocol, 103 metabolites were finally identified from a sample of S. coelicolor, which distribute in central metabolic pathways (glycolysis, pentose phosphate pathway and citrate cycle), amino acid, fatty acid, nucleotide metabolic pathways, etc. By comparing the temporal profiles of these metabolites, the amino acid and fatty acid metabolic pathways were found to stay at a high level during stationary phase, therefore, these pathways may play an important role during the transition between the primary and secondary metabolism. An optimized protocol of sample preparation was established and applied for metabolomics analysis of S. coelicolor, 103 metabolites were identified. The temporal profiles of metabolites reveal amino acid and fatty acid metabolic pathways may play an important role in the transition from primary to secondary metabolism in S. coelicolor.
Gas Chromatography-Mass Spectrometry ; Metabolic Networks and Pathways ; Metabolome ; Metabolomics ; methods ; Streptomyces coelicolor ; metabolism

Objective To describe the anatomical variants of infants’internal cerebral veins and their tributaries with MR suscep-tibility weighted imaging,and to evaluate the capability in the visualization of cerebral deep veins.Methods 80 healthy infants were enrolled in this study.All the brain images were obtained by a 3D gradient-echo sequence (Enhanced T2 ? weighted angiography-ES-WAN)on a 3.0T MR,which were reconstructed with minimal intensity projection.The septal vein,thalamostriate vein,internal cerebral vein and anterior caudate nucleus veins were evaluated.4 types (ⅠA,ⅠB,ⅡA,ⅡB)were classified based on their rela-tionship with the septal vein-internal cerebral vein junction and interventricular foramen,and 3 types (Ⅰ,Ⅱ,Ⅲ)based on the drainage patterns of the anterior caudate nucleus veins.Results The septal vein,thalamostriate vein and internal cerebral vein could be clearly and continuously visualized in 100% infants,and the visualization rate of the anterior caudate nucleus veins was 88.1%. TypeⅠA and TypeⅠ were most common in the two classification patterns.Conclusion Infants’internal cerebral veins and their tributaries are able to be clearly shown with the minimal intensity projection in susceptibility weighted imaging,which is a good method to evaluate the cerebral deep veins in infants.

Objective To understand the carrying status and drug resistance of streptococcus agalactiae in the genital tract of pregnant women.Methods The vaginal discharge from pregnant women was collected for conducting the bacterial culture.The iso-lated streptococcus agalactiae was performed the identification and the drugs susceptibility test by the VITEK 2 fully automatice bacterial identification analyzer.Results Among 1 042 samples,streptococcus agalactiae was isolated in 6 cases with the separation rate of 5.4%.The drug sensitivity test showed the Streptococcus agalactiae was highly sensitive to penicillin,cefazolin,vancomycin, linezolid and nitrofurantoin.The intermediate strains or drug resistant strains were not found.The sensitive rate to levofloxacin, moxifloxacin and clindamycin,erythromycin,tetracycline were 84.0%,84.0%,71.4%,39.3% and 8.9% respectively.Conclusion The carrying rate of streptococcus agalactiae in pregnant women is not high in our hospital.Penicillin and cefazolin should be as the drug of first choice for therapy and the clincal and laboratory nould payattention to detection of streptococcus agalactiae in pregnant women.