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Cheng L.-K.,Tianjin University of Science and Technology | Wang J.,Jilin University | Xu Q.-Y.,Tianjin University of Science and Technology | Zhao C.-G.,Tianjin University of Science and Technology | And 3 more authors.
World Journal of Microbiology and Biotechnology | Year: 2013

Optimum production of l-tryptophan by Escherichia coli depends on pH. Here, we established conditions for optimizing the production of l-tryptophan. The optimum pH range was 6. 5-7. 2, and pH was controlled using a three-stage strategy [pH 6. 5 (0-12 h), pH 6. 8 (12-24 h), and pH 7. 2 (24-38 h)]. Specifically, ammonium hydroxide was used to adjust pH during the initial 24 h, and potassium hydroxide and ammonium hydroxide (1:2, v/v) were used to adjust pH during 24-38 h. Under these conditions, NH4+ and K+ concentrations were kept below the threshold for inhibiting l-tryptophan production. Optimization was also accomplished using ratios (v/v) of glucose to alkali solutions equal to 4:1 (5-24 h) and 6:1 (24-38 h). The concentration of glucose and the pH were controlled by adjusting the pH automatically. Applying a pH-feedback feeding method, the steady-state concentration of glucose was maintained at approximately 0. 2 ± 0. 02 g/l, and acetic acid accumulated to a concentration of 1. 15 ± 0. 03 g/l, and the plasmid stability was 98 ± 0. 5 %. The final, optimized concentration of l-tryptophan was 43. 65 ± 0. 29 g/l from 52. 43 ± 0. 38 g/l dry cell weight. © 2012 Springer Science+Business Media Dordrecht.

Hao X.,China Agricultural University | Shen Z.,Shandong Binzhou Animal Science and Veterinary Medicine Institute | Wang J.,Shandong Binzhou Animal Science and Veterinary Medicine Institute | Zhang Q.,University of Manitoba | And 3 more authors.
Veterinary Journal | Year: 2013

Slightly acidic electrolyzed water (SAEW, pH 5.0-6.5) is a novel disinfectant with environmentally friendly broad spectrum microbial decontamination properties which could have significant utility on farm. Two of the most important pathogenic viruses in pigs are porcine reproductive and respiratory syndrome virus (PRRSV) and pseudorabies virus (PRV). The aim of this study was to evaluate the viricidal effectiveness of SAEW against PRRSV and PRV in vitro under different available chlorine concentrations (ACCs, 30, 50 and 70. mg/L), treatment times (5, 10 and 15. min) and temperatures (4, 20, 40 and 60 °C), respectively. SAEW had a strong viricidal activity against both PRRSV and PRV. This activity increased with increasing ACC, treatment time and temperature. PRRSV and PRV titres of 7.0log10TCID50/mL and 5.9log10TCID50/mL, respectively, were completely inactivated by SAEW at an ACC of ≥50mg/L for 10min even though SAEW had no negative effect on the host cells. SAEW thus shows promise as a disinfectant for use on pig farms to reduce the spread of both PRRSV and PRV, and to limit the morbidity associated with those viruses. © 2013 Elsevier Ltd.

Wang J.,Jilin University | Cong Y.,Jilin University | Yin R.,Jilin University | Feng N.,Academy of Military Medical Science | And 10 more authors.
Virus Research | Year: 2015

Newcastle disease virus (NDV) and Goose parvovirus (GPV) are considered to be two of the most important and widespread viruses infecting geese. In this study, we generated a recombinant rmNA-VP3, expressing GPV VP3 using a modified goose-origin NDV NA-1 by changing the multi-basic cleavage site motif RRQKR↓F of the F protein to the dibasic motif GRQGR↓L as that of the avirulent strain LaSota as a vaccine vector. Expression of the VP3 protein in rmNA-VP3 infected cells was detected by immunofluorescence and Western blot assay. The genetic stability was examined by serially passaging 10 times in 10-day-old embryonated SPF chicken eggs. Goslings were inoculated with rmNA-VP3 showed no apparent signs of disease and developed a strong GPV and NDV neutralizing antibodies response. This is the first study demonstrating that recombinant NDV has the potential to serve as bivalent live vaccine against Goose parvovirus and Newcastle disease virus infection in birds. © 2015 Elsevier B.V.

Mei J.-G.,Shandong Binzhou Animal Science and Veterinary Medicine Institute | Shen Z.-Q.,Shandong Binzhou Animal Science and Veterinary Medicine Institute | Zhuang J.-Q.,Shandong Binzhou Animal Science and Veterinary Medicine Institute
Chinese Journal of Biologicals | Year: 2010

Objective: To produce recombinant human tissue type plasminogen activator (rht-PA) by culture of recombinant CHO cells in 30 L packed bed bioreactor. Methods: The CHO cell strain for expression of rht-PA was resuscitated with IMDM containing 10% fetal bovine serum (FBS), then amplified and inoculated to a 30 L bioreactor under real-time monitoring by Biocommand Plus software. Serum-containing medium was used for culture of CHO cells at first then was changed into serum-free one. Perfusion culture mode was adopted in the whole course of culture, during which the glucose concentration in culture supernatant was determined every day, and expression level and bioactivity of rht-PA every other day. The expressed rht-PA was purified by Lysine-Sepharose 4B and Zn 2+-Sepharose 4B affinity chromatography and determined for specific activity, yield and purity. Results: The whole culture course lasted for 51 d, consisting of serum-containing culture for 6 d for cell growth and serum-free culture for 45 d for rht-PA expression. The perfusion rate was 46.7 L/d in average and 60 L/d at the most. About 2 100 L of culture supernatant was harvested. The expression level of rht-PA was 15.15 mg/L in average and 19.25 mg/L at the most. The mean bioactivity of rht-PA was about 8 000 IU/ml. The glucose consumed reached a peak value of 15.97 g/L·d on day 13 after serum-free culture for expression. The specific activity, yield and purity of purified rht-PA were 6 × 10 5 IU/mg, 63% and more than 99% respectively. Conclusion: A 30 L packed bed bioreactor was suitable for long-term continuous culture of recombinant CHO cells and high expression of rht-PA.

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