The yeast Sacchromyces cerevisiae is most widely used for producing bioethanol in alcoholic industry due to its higher ethanol yield and fermentation rate. However, the toxic effect of accumulated ethanol is one of the main factors, which limit high ethanol production. Thus, investigating the mechanisms of yeast ethanol tolerance will provide the basis for solving the industrial problem. This article reviewed the mechanisms of Sacchromyces cerevisiae ethanol tolerance focusing on its cell physiological behaviors, structure and biochemical composition, as well as its genetic basis.

A model for screening the yeast which can ferment xylose to produce the ethanol was constructed.An ethanol yeast was obtained using the lignocellulose as substrate production the ethanol.By malt extract medium pre-culturing,soil samples use the plate with xylose as sole carbon source as the primary screening,then finally screen by the potassium dichromate color-displaying method.A strain named Y2-3 was screened from the soil.Phenotypic analysis including morphology and physiology and biochemical characteristics and 26D1/D2 sequence analysis were carried out.Based on taxonomy results,the Y2-3 was identified as Pichia caribbica.The strain Y2-3 ferments using xylose as sole carbon source: biomass 23.5 g/L,xylose utilization rate 94.7 %,ethanol final yield 4.57 g/L;using mixture sugar:biomass 28.6 g/L,xylose utilization rate 94.2 %,glucose utilization rate 95.6%,ethanol final yield 20.6 g/L.Pichia caribbica is a yeast which can utilize xylose and mixture sugar as substrate.It established the foundation for further research fermentation of ethanol by yeast using lignocellulose.

Based on the principle of the pathway engineering, a novel pathway of producing glycerol was built in E. coli. The gpd1 gene encoding glycerol 3-phosphate dehydrogenase and the hor2 gene encoding glycerol 3-phosphatase were cloned from Saccharomyces cerevisiae, respectively. The two genes were inserted into expression vector pSE380 together. A recombinant plasmid pSE-gpd1-hor2 containing polycistron was constructed under the control of the strong trc promoter. Then it was transformed into E. coli BL21. The result showed the recombinant microorganism GxB-gh could convert glucose to glycerol directly. And the recombinant microorganism GxB-gh was incubated to produce glycerol from D-glucose in the fermentor. The maximal concentration of glycerol was 46.67g/L at 26h. Conversion rate of glucose was 42.87%. The study is about "green" producing glycerol by recombinant microorganism and is also useful for further working in recombining microorganism of producing 1,3-propanediol.
Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Fermentation ; Fungal Proteins ; biosynthesis ; genetics ; Genetic Engineering ; Glycerol ; metabolism ; Glycerolphosphate Dehydrogenase ; biosynthesis ; genetics ; Phosphoric Monoester Hydrolases ; biosynthesis ; genetics ; Saccharomyces cerevisiae ; enzymology ; genetics

1,3-propanediol (1,3-PD) is an important material for chemical industry, therefore, there is much interest in the production of 1,3-PD. The gene dhaT encoding 1, 3-propanediol dehydrogenase ( PDOR) of Citrobacter freundii was amplified by PCR. Sequence analysis of the similarity at the nucleotide and amino acid level between the gene encoding C. freundii PDOR and that of C. freundii ( U09771 ) were 78% and 90% , respectively. The recombinant plasmid pSE-dhaT was constructed by inserting dhaT gene into expression vector pSE380 and then transformed E. coli JM109. The recombinant strain was induced by IPTG to express dhaT. Further more the recombinant enzyme was purifed from recombinant E. coli by Ni-nitrilotriacetate affinity chromatography followed by Sephacral S-300 gel filtration. A single obvious protein about 42kDa could be obtained by the sodium dodecyl sulfate-polyacrylamide gel electrophoresis of recombinant enzyme. The purified enzyme was used to determined enzyme property on the substrate of propionaldehyde and 1, 3-PD. The optimal temperature and optimal pH of the purified enzyme were 37℃, 8.0 for reduction and 25℃, 10. 5 for oxidation, respectively; and the kinetic property of PDOR about Km and V max were 10. 05mmol/L, 37. 27?mol/min/mg for propionaldehyde and 1. 28mmol/L, 25. 55?mol/min/mg for 1,3-PD, respectively; The deduced dhaT gene product (388 amino acids) showed a specific reduction activity of 49. 50U/mg and oxidation activity of 79. 92U/ mg. There also have a putative iron-binding motif ( G-XX-H-X-X-A-H-X-X-G-X-X-X-X-X-P-H-G) as a fingerprint pattern in the recombinant enzyme, the motif is fully conserved among these 1, 3-propanediol dehydrogenase. It is beneficial to the researches of high producing 1, 3-propanediol by gene engineering strain.

OBJECTIVEThe choice of surgical approaches for salvage surgery based on the location and invasion of recurrent and residual lesions of nasopharyngeal carcinoma (NPC), surgical results, complications, and survival were assessed.

METHODSThirty-seven cases with recurrent and residual lesions of NPC underwent salvage surgery between March 1991 and January 2005 were analysed retrospectively. Of 37 patients, 23 were men and 14 women, with a median age of 46.5 years (26 - 57 years); 4 were at stage I, 10 at stage II, 14 at stage III, and 9 at stage IV; 5 cases were with cervical metastasis, including 3 cases of N1 and 2 cases N2. All recurrent and residual lesions of NPC were determined by biopsy. On the location and invasion of recurrent and residual lesions of NPC, 8 cases underwent endoscopic resection of lesions, 12 cases of the palate nasopharyngectomy, 5 cases of maxillary swing, 4 cases of maxillary swing plus prerenal approach, 2 cases of lateral rhinotomy plus coronal flap approach, and 6 cases transfacial plus nasal pyramid swing approach. Five cases with cervical metastasis received neck dissection in addition to the operations for recurrent and residual lesions of NPC. Postoperatively 31 cases received radiotherapy with dosage of 60 Gy, among them 15 cases with concurrent chemoradiation therapy, and 6 cases with clear surgical margin did not received radiotherapy or chemotherapy. The cases were followed up for 12 - 72 months, with a median of 45 months.

RESULTSTotal resection for the recurrent and residual lesions of NPC accounted for 91.8% (34/37) and subtotal resection for 8.2% (3/37). The accident of perioperative complications was 24.3% (9/37). The 3- and 5-year overall disease-free survival rates (DFSR) were 62.1% and 43.3%, respectively. The 3- and 5-year overall survival rates (OSR) were 72.9% and 51.3%, respectively. The 5 year DFSR of cases at stage I-IV were 100%, 40%, 28% and 11% (χ(2) = 10.0, P < 0.01), respectively. The 5 year OSR were 100%, 70%, 35% and 28% (χ(2) = 11.5, P < 0.01), respectively.

CONCLUSIONSSalvage surgery is a justified treatment for the recurrent and residual lesions of NPC, by which some patients with recurrent and residual lesions of NPC can be salvaged.

Adult ; Female ; Humans ; Male ; Middle Aged ; Nasopharyngeal Neoplasms ; pathology ; surgery ; Neoplasm Recurrence, Local ; surgery ; Neoplasm Staging ; Neoplasm, Residual ; Prognosis ; Retrospective Studies ; Salvage Therapy ; methods

Adult ; Aged ; CD4-Positive T-Lymphocytes ; Case-Control Studies ; Female ; Humans ; Interleukin-17 ; blood ; Liver Cirrhosis, Biliary ; blood ; immunology ; metabolism ; Male ; Middle Aged ; T-Lymphocytes, Helper-Inducer

OBJECTIVETo study the relationship between multidrug-resistant (MDR) expression in nasopharyngeal carcinoma (NPC) and its sensitivity to chemotherapy.

METHODSThe specimens of 23 NPC cases were studied by immunohistochemistry with monoclonal antibody of P-glycoprotein (P-gp), multidrug resistance relation protein (MRP), lung-resistance related protein (LRP), topoisomerase II (Topo II), thymidylate synthase (TS), glutathione-S-transferase (GST-pi). Among them, 20 specimens were taken from primary NPC lesion which were treated with two course of cisplatin (DDP) and 5-fluorouracil (5-FU), 3 specimens were taken from cervical lymph-node of recurrent NPC patients who were treated by radical dissection.

RESULTSVarious MDR parameters were expressed differently in 22 cases except for 1 clear cell carcinoma case. The difference was statistically significant (P < 0.05). However, there were no significant difference of MDR expression either among various carcinoma pathomorphology cell groups or among different clinical stage groups. Expression of LRP and TS were found in 10 and 14 cases respectively and the chemotherapy responders rates were 20% (2/10) and 28.5% (4/14) respectively. While the chemotherapy responders rates were 70% (7/10) and 5/6 in cases without expression. There was significant difference (P < 0.001, and P < 0.05).

CONCLUSIONThe NPC patients with LRP and TS expression may be less sensitive to chemotherapy with DDP + 5-FU.

ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; Adult ; Aged ; Cisplatin ; pharmacology ; therapeutic use ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Drug Screening Assays, Antitumor ; Female ; Fluorouracil ; pharmacology ; therapeutic use ; Glutathione S-Transferase pi ; genetics ; Humans ; Male ; Middle Aged ; Nasopharyngeal Neoplasms ; drug therapy ; genetics ; Thymidylate Synthase ; genetics ; Vault Ribonucleoprotein Particles ; genetics

OBJECTIVETo investigate the inhibitory effect of photodynamic therapy (PDT) in combination with paclitaxel (PCT) on proliferation in esophageal carcinoma Eca-109 cells line.

METHODSEca-109 cells were treated with PCT alone, HPD alone at different doses, or their combinations. For the combined treatments, the cells were exposed to PCT for 12 h followed by incubation with HPD at high, middle or low concentrations for 4 h. PDT was then performed on these treated cells and fluorescence microscopic observation was made before and after PDT. The cell survival was measured by MTT assay, and the cell apoptosis rate analyzed by flow cytometry after a 24-h cell incubation following PDT.

RESULTSThe fluorescence excitation of the cells was weakened after PDT. Combined treatments resulted in significantly lowered cell survival rate and increased cell apoptosis rates as compared to those of the control cells and the cells treated with PCT alone and low-dose HPD (P<0.01). Significant differences were also noted among the cells exposed to HPD at different concentrations (P<0.05).

CONCLUSIONPDT combined with PCT have significant synergetic effects in inhibiting the proliferation of human esophageal carcinoma cells and inducing their apoptosis in vitro.

Antineoplastic Agents, Phytogenic ; pharmacology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Esophageal Neoplasms ; pathology ; Humans ; Paclitaxel ; pharmacology ; Photochemotherapy

OBJECTIVETo study the resistant rate of hepatitis B virus (HBV) to ADV and the dynamic evolution of HBV in lamivudine (Lam)-resistant chronic hepatitis B (CHB) patients.

METHODSTwenty-three Lam-resistant CHB patients were assigned to a 10mg/d ADV monotherapy for 68-116 weeks. The baseline and different time point blood samples after ADV monotherapy were analyzed for ADV-resistant mutations using direct sequencing of PCR products; the evolution of HBV mutations was examined by clonal analysis of serial samples from one patient infected with ADV-associated resistant HBV strains.

RESULTSThe cumulative incidence of genotypic ADV resistance at weeks 48 and 96 was 4.3% and 10.5% respectively respectively. The evolution analysis of HBV mutant strains in an ADV-resistant CHB patient showed that the proportion of YMDD mutants gradually decreased with rtA181S mutants increasing over time after ADV monotherapy, and that rtA181S+N236T mutants became the predominant strains during prolonged ADV monotherapy. The addition of Lam to the ongoing ADV treatment had poorer antiviral response in the patient with rtA181S or rtA181S+N236T mutant infection; one clone with multi-drug resistant mutations was selected during Lam and ADV combination therapy.

CONCLUSIONIncreased risk of adefovir resistance and selection of multi-drug resistant mutations are associated with long-term ADV monotherapy in patients with Lam-resistant chronic hepatitis B.

Adenine ; analogs & derivatives ; therapeutic use ; Adult ; Antiviral Agents ; therapeutic use ; Drug Resistance, Viral ; Evolution, Molecular ; Female ; Hepatitis B virus ; classification ; drug effects ; genetics ; Hepatitis B, Chronic ; drug therapy ; virology ; Humans ; Lamivudine ; pharmacology ; Male ; Middle Aged ; Organophosphonates ; therapeutic use

Brown ducks carrying DHBV were widely used as hepatitis B animal model in the research of the activity and toxicity of anti-HBV dugs. Studies showed that the ratio of DHBV carriers in the brown ducks in Guilin region was relatively high. Nevertheless, the characters of the DHBV genome of Guilin brown duck remain unknown. Here we report the cloning of the genome of Guilin brown duck DHBV and the sequence analysis of the genome. The full length of the DHBV genome of Guilin brown duck was 3 027bp. Analysis using ORF finder found that there was an ORF for an unknown peptide other than S-ORF, PORF and C-ORF in the genome of the DHBV. Vector NTI 8. 0 analysis revealed that the unknown peptide contained a motif which binded to HLA * 0201. Aligning with the DHBV sequences from different countries and regions indicated that there were no obvious differences of regional distribution among the sequences. A fluorescence quantitative PCR for detecting DHBV was establishment based on the recombinant plasmid pGEM-DHBV-S constructed. This study laid the groundwork for using Guilin brown duck as a hepatitis B animal model.
Animals ; Base Sequence ; China ; epidemiology ; Cloning, Molecular ; Ducks ; Genome, Viral ; Hepadnaviridae Infections ; diagnosis ; veterinary ; virology ; Hepatitis B Virus, Duck ; classification ; genetics ; isolation & purification ; Molecular Sequence Data ; Phylogeny ; Polymerase Chain Reaction ; methods ; Poultry Diseases ; diagnosis ; virology