Microbial lipases have been widely used in traditional industries,as well as in emerging biocatalysed areas owing to their ability to catalyze a variety of reactions in aqueous and non-aqueous media.Therefore,it is very important to enhance amount of lipase production by recombinant overexpression for meeting market demand.A critical review of main and novel strategies which have been employed for recombinant expression of microbial lipases are presented,including codon optimization,fusion and co-expression,dual expression system based on hybrid promoters,homologous overexpression,cell surface displaying and high-throughput screening based on gene library of expression.These new technologies are gradually coming to the forefront in the recombinant expression of lipase,especially for cell surface displaying and high-throughput screening based on gene library of expression.Meanwhile,several recombinant expressions for representative microbial lipases were also introduced and discussed,which are available for consultation when attempting to overexpress any lipases by scientists and industrialists.

Objective To investigate the transactivating effect of hepatitis C virus(HCV)core protein on inducible nitric oxide synthase(iNOS)gene promoter and the molecular biological mecha- nisms of HCV pathogenesis.Methods Polymerase chain reaction(PCR)technique was employed to amplify the sequence of iNOS promoter by using HepG2 genomic DNA as template,and the product was cloned into pGEM-T vector.The iNOSp gene was cut from T-iNOSp by KpnⅠand XhoⅠ,and then was cloned into pCAT3-Basic,the constructed vector was named as pCAT3-iNOSp,pCAT3-iN- OSp was transfected into the LO_2 cell line.LO_2 cell was also cotransfected with pcDNA3.1(-)-core and pCAT3-iNOSp by FuGENE 6 transfection reagents.The LO_2 cells transfected with pCAT3-Basic was used as negative control.The activity of CAT in LO_2 cells was detected by an ELISA kit after 48 hours,which reflected the transactivating function of HCV core protein to iNOS gene promoter.Re- sults The expressive vector pcDNA3.1(-)-core and report vector pCAT3-iNOSp had been construc- ted and confirmed by restriction enzyme digestion and sequencing.The expression of CAT in LO_2 cells transfected with pCAT3-iNOSp and peDNA3,1(-)-core was 11 times as higher as that of pCAT3-bas- ic,and 6 times as higher as that of pCAT3-iNOSp.Conclusion It is suggested that HCV core protein can transactivate iNOS gene promoter.

Purpose:To study the effect of inhibition of telomerase activity and cell cycle by transcriptase telomerase inhibitors (3′ azido 3′ deoxythymidine, AZT) on squamous cell carcinoma of tongue in vitro.Methods:Human squmous cell carcinoma of tongue cell line Tca8113 was used as target cell. Telomerase activity was determined by TRAP PCR ELISA in untreated and treated Tca8113 by AZT, cell cycle phases were analyzed by flow cytometry. Results:Telomerase activity of Tca8113 was significantly inhibited when treated with AZT, and the effect of inhibition was dose dependant (rate of telomerase activity treated with AZT in 0.3, 0.6, 1.0, 1.5mol 10 -1 was 0.69 0.03, 0.61 0.08, 0.53 0.11, 0.50 0.02 respectively, rate of telomerase activity treated without AZT was 0.76 0.06). Cell cycle of treated Tca8113 was changed with marked increase in G 2 /M phase compared with untreated Tca8113 (62.8% vs 19.7%, P

Objective:To develop an HPLC method for the determination of capsaicine, dihydrocapsaicin, imperatorin and isoim-peratorin in Guanjie Jietong plasters. Methods:A CAPCELL PAK C18 column was used as the chromatographic column, and the flow rate was 0. 8 ml·min-1. The mobile phase A consisted of methanol-acetonitrile(1∶1), the mobile phase B was of 1% phosphoric acid. The detection wavelength was 280 nm and 254 nm, respectively. Results:There was a good linear relationship between the con-centrations and the peak areas within the range of 0. 298-5. 956μg(r=0. 9998) for capsaicine, 0. 152-3. 044μg(r=0. 999 6) for di-hydrocapsaicin, 0. 018-0. 352 μg(r=0. 999 5) for imperatorin and 0. 010-0. 204 μg(r=0. 999 3) for isoimperatorin. The average re-covery was 97. 8%(RSD=1. 02%), 97. 0%(RSD=0. 76%), 96. 6%(RSD=0. 65%) and 98. 1%(RSD=1. 35%), respectively. Conclusion:The method is convenient, accurate, sensitive and repeatable, which can be used in the determination of capsaicine, di-hydrocapsaicin, imperatorin and isoimperatorin in Guanjie Jietong plasters.

The fermentation conditions of lipase production by Geotrichum candidum Y162 were optimized. Initially, the most suitable carbon olive oil, nitrogen source soybean flour and NH4Cl, salt BaCl2 and MgCl2 were selected according to single factorial experiments respectively. Based on the result, screening methodology Plackett-Burman design was used to evaluate the effects of twelve factors related to lipase production and three statistically significant factors olive oil, BaCl2 and NH4Cl were selected. The path of steepest ascent was used to approach the optimal region of lipase production subsequently. Then, the optimal combined concentration for maximum enzyme activity were further optimized by response surface methodology and determined as follows: olive oil 2.35%, BaCl2 0.36%,and NH4Cl 4.69%.The optimization of culture conditions of G.candidum Y162 led to a 2.25-fold increase in lipase production relative to initial result 14.16 U/ml, which indicate that single factor in combination with response surface methodology is an effective method for optimization of lipase production conditions by G.candidum Y162.

Adult ; Aged ; Humans ; Middle Aged ; Pneumoconiosis ; diagnostic imaging ; Radiography, Thoracic ; Retrospective Studies

Human bocavirus (HBoV) 1-4 have been detected both in respiratory and stool samples since the first HBoV was discovered in 2005. HBoV-1 is mostly associated with respiratory infection, while HBoV 2-4 are usually associated with intestinal tract infection. A variety of signs and symptoms have been described in patients with HBoV infection, including cough, wheezing, pneumonia, and diarrhea, but the research on pathogenic mechanism of HBoV is limited because HBoV cannot be cultured in vitro due to the lack of appropriate host cells. Three-dimensional epithelial cell culture, reverse genetics, and viral metagenomics are identified as novel tools that may promote the research on pathogenic mechanism of HBoV and the discovery of new viruses. This review summaries currently available diagnostic approaches such as electron microscopy, cell culture, PCR, and immunoassay in order to provide a method reference for indepth research on HBoV.
Animals ; Human bocavirus ; genetics ; growth & development ; isolation & purification ; pathogenicity ; Humans ; Parvoviridae Infections ; diagnosis ; virology ; Viral Proteins ; genetics ; metabolism ; Virology ; methods ; Virulence ; Virus Cultivation

Ear Neoplasms ; Ear, Middle ; pathology ; Female ; Humans ; Mastoid ; pathology ; Melanoma ; Middle Aged

Electrocardiography ; Humans ; Long QT Syndrome

Objective To study the function of cyclooxygenase 2(COX 2) and its specific inhibitor NS 398 on the cell growth and invasion ability of urothelial carcinoma cell line EJ. Methods The cox 2 cDNA was transfected into the urothelial carcinoma cell line EJ and a cell line EJ COX 2 which highly expressed cox 2 gene permanently was gained. The cell growth rate before and after transfection was observed. Then at various concentrations of NS 398, the invasion ability was detected by Boyden Chamber and expression levels of uPA by RT PCR and Western blot. Results The EJ COX 2 cell line grew more rapidly and had a stronger invasion ability than EJ and its uPA expression increased significantly. NS 398 could dose dependently inhibit the expressions of COX 2 and uPA and the invasiveness of EJ COX 2 cell. Conclusion COX 2 can stimulate the growth of urothelial cell line EJ and promote its invasion ability by stimulating the expression of uPA.