Statistical optimization of medium composition for aspergiolide A production by marine-derived fungus Aspergillus glaucus.

Statistical methodologies were employed to optimize submerged culture medium for the production of a novel antineoplastic compound aspergiolide A by a marine-derived fungus Aspergillus glaucus HB1-19 for the first time. Orthogonal design was preformed to determine the initial composition. Then Plackett-Burman design was applied to evaluate the influence of related nutrients, and yeast extract paste, soybean powder and sodium glutamate were confirmed as critical factors in the medium.

[Refolding of the fusion protein of recombinant enterokinase light chain rEKL].

The fusion protein of enterokinase light chain, DsbA-rEKL, was expressed mainly in inclusion body in E. coli. The recombinant bacteria was fermented to high density, with high expression of the fusion protein.

[Detection of intracellular reactive oxygen species by flow cytometry in Pichia pastoris fermentation].

In Pichia pastoris fermentation, methanol was oxidized into carbon oxide and produced a byproduct H2 O2, one of the partially reduced forms of molecular oxygen known as reactive oxygen species (ROS) . ROS are highly damaging towards cellular constituents. Flow cytometry (FCM) is an excellent method that permits the rapid, optical analysis of individual cells and has many advantages over conventional cytometry.

[Metabolism of recombinant CHO-GS cell reducing of toxic effect of ammonia].

The toxic effect of ammonia on rCHO-GS cell decreased obviously due to the transfection of GS system in serum-free culture. The maximum cell density, 15.6 x 10(5) cells/mL was obtained in the culture with 1.

[Fermentation behaviors of recombinant Pichia pastoris under inhibited methanol concentration].

Chemostat culture was performed to characterize the growth, substrate consumption and the hirudin production, and to disclose their interrelations in the fermentation of recombinant Pichia pastoris. The Andrew substrate-inhibited growth model is more suitable than Monod model to simulate the growth of Pichia pastoris on methanol. Therefore, two stationary states can be obtained in the continuous culture at a certain dilution rate because of the substrate inhibition on cell growth.

Purification and identification of recombinant hirudin and its degradation products expressed in Pichia pastoris.

The purification and identification of recombinant hirudin (r-hirudin) (rHV2-Lys47) and its several C-terminal proteolytic degradation derivatives, produced by Pichia pastoris, were described. The high-purity rHV2-Lys47 of above 99% and its three degradation products were obtained by a straightforward two-step chromatography procedure, a combination of cation exchange and reverse phase chromatography, with a recovery yield of 74% for hirudin. The purified rHV2 had the predicted N-terminal amino acid sequence and the derivatives were the degradation products of hirudin, short of one to three amino acid residues at C-terminal.

[Microbial transformation of sinenxan A, a rich constituent in callus cultures of Taxus].

Aim: To study the microbial transformation of sinenxan A.

Methods: Choose two strains of Fungi (Mucor spinosus AS 3.3450 and Cunninghamella echinulata AS 3.

DNA sequencing of a plasmid with virulence from marine fish pathogen Vibrio anguillarum.

DNA sequence of a plasmid pEIB1 associated with virulence from the marine fish pathogen Vibrio anguillarum was determined using the methods of restriction endonuclease digestion, subcloning, and primer walking. The whole length of obtained pEIB1 DNA sequence was 66 164 bp, and the overall G+C content of DNA sequence is 42.7%.

Estimation of Chinese hamster ovary cell density in packed-bed bioreactor by lactate production rate.

A method is described for estimating recombinant Chinese hamster ovary (rCHO) cell density in a packed-bed bioreactor by lactate production rate. The lactate production rate, which depended on both the cell numbers and cell growth rate, was modeled by segregating the cell population into two parts: one growing at a maximum specific growth rate and another non-growing. The individual cell in each part had the same lactate production rate.

Promoter activities in the baculovirus envelope glycoprotein gp64 gene.

Baculovirus GP64 envelope glycoprotein is a specific major component of the envelope of the budded virus and is involved in virus entry into the host cells by endocytosis. For promoter activity analysis in the baculovirus gp64 gene, two DNA fragments containing 437 and 439 bp upstream of 5' ends of the BmNPV and AcMNPV gp64 ORF were amplified by polymerase chain reaction and cloned, respectively. The sequence analysis indicated that two gp64 genes have both early (CAGT) and late (A/GTAAG) transcriptional start sites.

Microbial metabolism of artemisinin by Mucor polymorphosporus and Aspergillus niger.

Artemisinin (1) was transformed by Mucor polymorphosporus and Aspergillus niger. Five products were identified as 9beta-hydroxyartemisinin (2), 3beta-hydroxyartemisinin (3), deoxyartemisinin (4), 3beta-hydroxydeoxyartemisinin (5), and 1alpha-hydroxydeoxyartemisinin (6). Products 2, 3, and 6 are new compounds.

[Effects of different methanol feeding strategy on hirudin production in high-density fermentation by recombinant Pichia pastoris].

Four different methanol feeding modes were evaluated in the hirudin production in high-density fermentation by Pichia pastoris. It was difficult to avoid methanol excessive in the broth with the feeding strategy only based on DO level. On the other hand, the fluctuation in methanol concentration was observed with methanol feeding strategy by off-line gas chromatography.