A 1.8 kb HinfI fragment carrying the chitinase gene(chiA) from Serratia marcescens was isolated from the plasmid pLCHIA and was inserted into the SmaI site downstream of the strong Ptac promoter in the expression vector pKK223-3,yielding the plasmid pKChiA.The 2.1 kb PtacChiA fusion fragment was excised by BamHI from the plasmid pKChiA and was inserted into the single BamHI site present within the plasmid pMC71A,thus generating the plasmid pMChiA.High levels of chitinase were produced by the E.coli strains HB101 and JM105 carrying the plasmid pKChiA or the plasmid pMChiA,with the amount of production were higher 1～3 fold than that produced by the strains carrying the plasmid pLCHIA.

Air Pollutants, Occupational ; analysis ; Solvents ; analysis

Objective To investigate the effects of total anthraquinone in rheum on aquaporin 2 and aquaporin 4 expression in rat kidney and explore its diuresis mechanism.Methods Thirty-two SD rats were randomly divided into control group,low dose group,medium dose group and high dose group.Total anthraquinone in rheum was administered to rats at different doses.Urinary volume of 24 h,Na+ concentration and osmolality were detected.Rats were sacrificed 5 days later.Blood samples were taken from the abdominal aorta to detect blood biochemical indicators. Kidneys of rats were removed to detect AQP2, AQP4 expression through immunohistochemistry,Western blot,and RT-PCR. Results Compared with control group,there were significantly increased 24 h urine output of rats in medium and high dose group[(16.21±1.96),(18.16±1.8) ml vs(13.85±1.25)ml,P＜0.05].24 h urine output in low-dose group did not change significantly.AQP2 protein and mRNA expression were significantly decreased in rats'kidneys of medium and high dose group (P＜0.01),The AQP4 protein and mRNA expression was significantly down-regulated in high dose group (P＜0.01).In medium does group,the AQP4 protein expression was down-regulated (P＜0.01),without significant decrease in the mRNA expression.Protein and mRNA expression of AQP2 and AQP4 did not significantly change in low dose group.Conclusion Total anthraquinone in rheum can reduce the expression level of AQP2 and AQP4 in rat kidney,which is probably one mechanism of diuresis caused by rheum.

Traditional herbal medicines, Panax ginseng, Panax quinquefolium and Panax notoginseng, attract our attention for their extensive and powerful pharmacological activities. Ginsenosides are the main active constituents of these medicinal herbs. The related glycosyltransferases involved in ginsenoside biosynthesis are the key enzymes which catalyze the last important step. Modification of ginsenoside aglycones by glycosyltransferases produces the complexity and diversity of ginsenosides, which have more extensive pharmacological activity. At present, ginsenoside aglycones and compound K have been obtained by synthetic biology. As the last step of ginsenoside biosynthesis, glycosylation of ginsenoside aglycones has been studied intensively in recent years. This review summarizes the basic strategies and research advances in studies on glycosyltransferases involved in ginsenoside biosynthesis, which is expected to lay the theoretical foundation for the in-depth research of biosynthetic pathway of ginsenosides and their production by synthetic biology.
Biosynthetic Pathways ; Ginsenosides ; biosynthesis ; Glycosyltransferases ; metabolism ; Panax ; chemistry ; Plants, Medicinal ; chemistry ; Synthetic Biology

According to morphological and microscopic characteristics, a high-efficient decolorizing fungus, the strain Asaw117, was identified as Aspergillus awamori. Selecting eight different dyes from Azo dyes, anthraquinones dyes and oxygen Quinones dyes, the decolorizing assays of various dyes showed that the strain Asaw 117 was the highest decolorizing potential to 0.1 g/L Vat Blue RSN, the discoloration rate up to 100 percent. Comparing to different kinds of medium and several of carbon and nitrogen sources, the strain had the best decolorizing efficient although grew slower in the Czapek medium, otherwise, grew quicker and decolorizing efficient lower in the PDF medium. It could use Vat Blue RSN as a nitrogen source, but not as a carbon source. The medium composing of saccharose and ammonium nitrate as carbon and nitrogen sources was decolorizing potential markedly during different combinations of carbon and nitrogen sources. So the strain has good potential for the dyeing wastewater treatment

[Summary]_ Transient neonatal diabetes mellitus is a kind of rare special types of diabetes. It should be distinguished from type 1 diabetes. Genetic analysis can be used to define the subtype of neonatal diabetes mellitus, which helps us to select the most appropriate treatment and to predict the disease recurrence. Sulfonylureas is able to improve insulin secretion in most patients with transient neonatal diabetes mellitus and provide effective glycemic control. A case of transient neonatal diabetes mellitus is reported in order to call attention to the diagnosis and treatment of this disease.

Aged ; Brain Abscess ; etiology ; Brain Neoplasms ; complications ; secondary ; Carcinoma, Squamous Cell ; complications ; secondary ; Humans ; Male ; Neoplasms, Unknown Primary ; Parietal Lobe

OBJECTIVETo study the relationship of TCM syndrome type of gastric mucosal epithelial growth factor (EGF), vascular endothelial growth factor (VEGF) and proliferative cell nuclear antigen (PCNA) in patients with chronic atrophic gastritis (CAG) for exploring the essence of TCM type and providing a theoretical basis of clinical treatment.

METHODSTCM syndrome type of 200 patients with diagnosis of CAG confirmed by fibro-gastroscope and pathological examination were differentially classified, and the expressions of EGF, VEGF and PCNA in different types were determined using immunohistochemistry.

RESULTSPatients were differentiated as Pi-Wei deficiency type (Type I ) in 72; Gan-Wei disharmony type (Type II ) in 43; Pi-deficiency with qi stagnation type (Type III) in 32; Wei-yin deficiency type (Type IV) in 24; Pi-Wei damp-heat type (Type V) in 14; and Wei-collateral stasis obstruction type (Type VI) in 5. The difference of PCNA expression level between Type II with Type I , III and IV was significant (P < 0.05). No significant difference in expression levels of EGF and VEGF was found among the 6 types (P > 0.05).

CONCLUSIONType I and II were the dominant TCM syndrome types in CAG patients; the high expression of PCNA might be a diagnostic evidence for Gan-Wei disharmony syndrome.

Adult ; Aged ; Aged, 80 and over ; Diagnosis, Differential ; Endothelial Growth Factors ; biosynthesis ; Female ; Gastric Mucosa ; metabolism ; pathology ; Gastritis, Atrophic ; diagnosis ; metabolism ; Humans ; Immunohistochemistry ; Male ; Medicine, Chinese Traditional ; Middle Aged ; Proliferating Cell Nuclear Antigen ; biosynthesis ; Syndrome ; Vascular Endothelial Growth Factor A ; biosynthesis

Lanosterol synthase is encoded by the erg7 gene and catalyzes the cyclization of 2, 3-oxidosqualene, which is a rate-limiting step of the inherent mevalonate (MVA)/ergosterol metabolic pathway in Saccharomyces cerevisiae. The intermediate 2, 3-oxidosqualene is also the precursor of triterpenoids. Therefore, the cyclization of 2, 3-oxidosqualene is the key branch point of ergosterol and triterpenoids biosynthesis. Down-regulation of 2, 3-oxidosqualene metabolic flux to ergosterol in S. cerevisiae may redirect the metabolic flux toward the triterpenoid synthetic pathway reconstructed by the synthetic biology approach. To construct erg7 knockout cassette harboring the loxP-Marker-loxP element, long primers were designed, which were homologous to the sequences of both erg7 ORF and plasmid pUG66. The cassette was transformed into diploid wild strain INVSc1 by LiAc/SS Carrier DNA/PEG method and then erg7 gene haploid deficient mutant was obtained by homologous recombination. The results of semiquantitative PCR and real-time quantitative PCR revealed that erg7 expression level in erg7 gene haploid deficient mutant is one time lower than that in wild strain. The results of TLC and HPLC showed that the ergosterol content in deficient mutant decreased to 42% of that in wild strain.
Chromatography, High Pressure Liquid ; DNA Primers ; Down-Regulation ; Ergosterol ; metabolism ; Haploidy ; Intramolecular Transferases ; genetics ; Polymerase Chain Reaction ; Saccharomyces cerevisiae ; genetics ; Squalene ; analogs & derivatives ; metabolism

Pichia pastoris is one of the most important systems used in the field of molecular biology for the expression of recombinant proteins. The system has advantages of high expression, high stability, high secretion, easy high-density fermentation and low cost. Many factors affect the expression of recombinant protein, such as gene copy number, codon usage preference, type of promoter, molecular chaperones, glycosylation, signal peptide and fermentation process. In this review, research advances of the above aspects are summarized, which lay a foundation for improving the expression of recombinant proteins in P. pastoris.
Fermentation ; Gene Dosage ; Glycosylation ; Molecular Chaperones ; Pichia ; metabolism ; Promoter Regions, Genetic ; Protein Sorting Signals ; Recombinant Proteins ; biosynthesis