Cloning of Laccase Gene from Coriolus Versicolor and Optimization of Culture Conditions for Lcc1 Expression in Pichia Pastoris

2011 ◽  
Vol 236-238 ◽  
pp. 1039-1044
Author(s):  
Li Yan He ◽  
Gui Bin Wang ◽  
Fu Liang Cao ◽  
Lin Guo Zhao ◽  
Yong Xin Ji
Keyword(s):  
Pichia Pastoris ◽  
Expression Vector ◽  
Culture Medium ◽  
Culture Conditions ◽  
Coriolus Versicolor ◽  
Laccase Production ◽  
Accession Number ◽  
Rt Pcr ◽  
Initial Ph ◽  
Native Signal Peptide

A laccase cDNA lcc1 (GenBank accession number HM137002), without native signal peptide, was cloned by RT-PCR from total RNA of Coriolus versicolor. Recombination expression vector pPICZαA-lcc1 was constructed and transformed into Pichia pastoris KM71H after lineared. Recombination laccase was expressed at a higher level. Single factors of fermentation conditions of Pichia pastoris KM71H for laccase production were optimized. The results showed optimal culture conditions were as follows: medium initial pH 7.5, Cu2+ concentration 0.5mmol/L, methanol additive amount 1.0% and shaker rotate speed 210r/min. Furthermore, induction at low temperature was more suitable for lcc1 secretion. And addition of appropriate amount peptone and tyrosine in culture medium could enhanced lcc1 yields and reduce its degradation.

Study on Optimization of Fermentation Condition for Nitrilase-Producer Escherichia Coli BL21 (DE3)/pET-Nit

2011 ◽  
Vol 175-176 ◽  
pp. 192-196 ◽  
Author(s):  
Li Li Feng ◽  
Jian Fei Zhang ◽  
Hui Luo ◽  
Zheng Li ◽  
Hong Jie Zhang
Keyword(s):  
Escherichia Coli ◽  
Recombinant Strain ◽  
Culture Medium ◽  
Culture Conditions ◽  
Fermentation Condition ◽  
Hydrogen Phosphate ◽  
Strain Bl21 ◽  
Initial Ph ◽  
Potassium Hydrogen ◽  
Optimization Of Fermentation

The paper concentrated on the optimization of the recombinant strain BL21 (DE3)-PE7-Nit. The component of culture medium and the culture conditions were optimized. The optimized medium was: yeast extract 10 g/l, L-glutamate sodium 8 g/l, MgSO4.7H2O 0.7 g/l, Isopropyl-β-D-thiogalactopyranoside 0.3 mmol/L, potassium hydrogen phosphate 0.5 g / L, phosphate Potassium 0.5 g / L and the culture condition was: initial pH 7.0, inoculum 2%. The result showed that the activity of nitrilase prepared with these conditions increased by 130.37 % through optimization.

Flocculating Activity Optimization of Lactobacillus paracasei subsp.paracasei L1 in Corn Liquid

2013 ◽  
Vol 850-851 ◽  
pp. 1172-1175
Author(s):  
Xiao Na Li ◽  
Xin Hua Li ◽  
Li Li Zhang ◽  
Jing Mu
Keyword(s):  
Culture Medium ◽  
Corn Starch ◽  
Culture Conditions ◽  
Lactobacillus Paracasei ◽  
Analysis Method ◽  
Medium Ph ◽  
Inoculum Concentration ◽  
Flocculating Activity ◽  
Initial Ph ◽  
Surface Analysis Method

In order to improve the production process of corn starch and accelerate the separation of corn starch, flocculating activity of lactobacillus paracasei subsp.paracasei L1 isolated from natural fermentation of sweet potato acid liquor in corn liquid was optimized. The main influencing factors were determined by Plackett-Burman experimental design and the flocculating activity was optimized by response surface analysis method. The optimum culture conditions was: inoculum concentration 18%, lactose addition 0.2%, corn steeping time 0h, culture temperature 30°C, initial pH of the culture medium pH 6.5, culture time 36h, yeast extract addition 1%,concentration of corn thick liquid 1:3(w/v).The theoretical value of the flocculating activity was 0.1906 mg/ml and the verified value was 0.1854 mg/ml.

Adsorption Applications of Bacillus sp. Biomass for Decolorization of Soluble Reactive Dyes Contaminated Solutions

2011 ◽  
Vol 71-78 ◽  
pp. 2294-2297
Author(s):  
Yan Li Mao ◽  
Yun Yu ◽  
Shi Tian Luo ◽  
Xiao Lin Lv
Keyword(s):  
Culture Medium ◽  
Reactive Dyes ◽  
Culture Conditions ◽  
Inoculum Size ◽  
Bacillus Sp ◽  
Light Yellow ◽  
Molasses Wastewater ◽  
Decolorization Efficiency ◽  
Flocculating Activity ◽  
Initial Ph

The optimal conditions of production of biopolymer by the culture of Bacillus sp.were examined, using molasses wastewater to replace glucose as carbon source and energy source in the culture medium. Results showed that the COD concentration in molasses wastewater favorable for the growth of the Bacillus sp.was 4500 mg•L-1, and inoculum size of 6%(v/v), 28◦C, initial pH 7.0 and shaking speed of 160 r•min-1, under the optimal culture conditions, the highest flocculating activity achieved for Kaolin suspension was 97.5% and 4.0 g biomass /L broth was obtained. The Bacillus sp. biomass was effective in flocculating some soluble reactive dyes in aqueous solution, reactive Light-Yellow K-4G and reactive Turquoise Blue KN-G with a decolorization efficiency of 98.5 and 92.5%, respectively, using 20 mL of the flocculant in 500 mL of 100 mg•L-1 dye solution.

Optimization of Removal Conditions of Soluble Reactive Dyes from Aqueous Solutions by Bacillus sp. Biomass

2012 ◽  
Vol 430-432 ◽  
pp. 395-398
Author(s):  
Gai Xia Fang ◽  
Li Hua Chen
Keyword(s):  
Energy Source ◽  
Culture Medium ◽  
Reactive Dyes ◽  
Culture Conditions ◽  
Inoculum Size ◽  
Bacillus Sp ◽  
Optimal Conditions ◽  
Light Yellow ◽  
Decolorization Efficiency ◽  
Initial Ph

The optimal conditions of by the culture of Bacillus sp.were examined, using beer wastewater to replace glucose as carbon source and energy source in the culture medium. Results showed that the COD concentration in beer wastewater favorable for the growth of the Bacillus sp.was 5000 mg•L-1, and inoculum size of 5%(v/v), 30◦C, initial pH 7.0 and shaking speed of 150 r•min-1, under the optimal culture conditions, the Bacillus sp. biomass was effective in flocculating some soluble reactive dyes in aqueous solution, reactive Light-Yellow K-4G and reactive Turquoise Blue KN-G with a decolorization efficiency of 93.8 and 95.5%, respectively, using 20 mL of the culture medium in 500 mL of 100 mg•L-1 dye solution.

Biosorption Applications of Flavobacterium sp. Biomass for Decolorization of Soluble Reactive Light-Yellow K-4G Solutions

2011 ◽  
Vol 322 ◽  
pp. 440-443
Author(s):  
Kui Zhang ◽  
Hong Qiao Xing ◽  
Yao Qing Wang
Keyword(s):  
Culture Medium ◽  
Reactive Dyes ◽  
Culture Conditions ◽  
Inoculum Size ◽  
Optimal Conditions ◽  
Light Yellow ◽  
Molasses Wastewater ◽  
Decolorization Efficiency ◽  
Flocculating Activity ◽  
Initial Ph

The optimal conditions of production of flocculant by the culture of Flavobacterium sp. were examined, using molasses wastewater to replace glucose as carbon source and energy source in the culture medium. Results showed that the COD concentration in molasses wastewater favorable for the growth of the Flavobacterium sp.was 5000 mg•L-1, and inoculum size of 5%(v/v), 30°C, initial pH 7.0 and shaking speed of 150 r•min-1, under the optimal culture conditions, the highest flocculating activity achieved for Kaolin suspension was 95.0% and 4.0 g biomass /L broth was obtained. The Flavobacterium sp. biomass was effective in flocculating some soluble reactive dyes in aqueous solution, reactive Light-Yellow K-4G with a decolorization efficiency of 98.5, using 20 mL of the flocculant in 500 mL of 100 mg•L-1 dye solution.

Production of a Novel Biopolymer by Culture of Bacillus cereus B-11 Using Molasses Wastewater and its Use for Dye Removal

2011 ◽  
Vol 230-232 ◽  
pp. 1119-1122 ◽  
Author(s):  
Yan Li Mao ◽  
Chang Xun Tian ◽  
Ji Wei Zhu ◽  
Tai Zhi Zhang ◽  
Lv Bao Tong
Keyword(s):  
Bacillus Cereus ◽  
Culture Medium ◽  
Dye Removal ◽  
Culture Conditions ◽  
Inoculum Size ◽  
Light Yellow ◽  
Molasses Wastewater ◽  
Decolorization Efficiency ◽  
Flocculating Activity ◽  
Initial Ph

The optimal conditions of production of biopolymer by the culture of Bacillus cereus B-11 were examined, using molasses wastewater to replace glucose as carbon source and energy source in the culture medium. Results showed that the COD concentration in molasses wastewater favorable for the production of the biopolymer was 5000 mg•L-1, and inoculum size of 5%(v/v), 28◦C, initial pH 7.0 and shaking speed of 150 r•min-1, under the optimal culture conditions, the highest flocculating activity achieved for Kaolin suspension was 98.5% and 3.55 g biopolymer /L broth was obtained. The biopolymer was effective in flocculating some soluble reactive dyes in aqueous solution, reactive Light-Yellow K-4G and reactive Turquoise Blue KN-G with a decolorization efficiency of 97.5 and 94.7%, respectively,using 25 mL of the flocculant in 500 mL of 100 mg•L-1 dye solution.

Cloning and Expression of Recombinant Human Insulin Gene in Pichia pastoris

2019 ◽  
Vol 10 (01) ◽  
Author(s):  
Rafid A. Abdulkareem
Keyword(s):  
Pichia Pastoris ◽  
Human Insulin ◽  
Expression Vector ◽  
Expression System ◽  
Insulin Gene ◽  
Cloning And Expression ◽  
Pcr Analysis ◽  
Major Band ◽  
Human Insulin Gene ◽  
Pichia Pastoris Expression System

The main goal of the current study was cloning and expression of the human insulin gene in Pichia pastoris expression system, using genetic engineering techniques and its treatment application. Total RNA was purified from fresh normal human pancreatic tissue. RNA of good quality was chosen to obtain a first single strand cDNA. Human preproinsulin gene was amplified from cDNA strand, by using two sets of specific primers contain EcoR1 and Notl restriction sites. The amplified preproinsulin gene fragment was double digested with EcoRI and Not 1 restriction enzymes, then inserted into pPIC9K expression vector. The new pPIC9K-hpi constructive expression vector was transformed by the heat-shock method into the E.coli DH5α competent cells. pPic9k –hpi, which was propagated in the positive transformant E. coli cells, was isolated from cells and then linearised by restriction enzyme SalI, then transformed into Pichia pastoris GS115 using electroporation method. Genomic DNA of His+ transformants cell was extracted and used as a template for PCR analysis. The results showed, that the pPic9k – hpi was successfully integrated into the P. pastoris genome, for selected His+ transformants clones on the anticipated band at 330 bp, which is corresponded to the theoretical molecular size of the human insulin gene. To follow the insulin expression in transformans, Tricine–SDS gel electrophoresis and Western blot analysis were conducted. The results showed a successful expression of recombinant protein was detected by the presence of a single major band with about (5.8 KDa) on the gel. These bands correspond well with the size of human insulin with the theoretical molecular weight (5.8 KDa).

scholarly journals Sensitive and Selective Culture Medium for Detection of Environmental Clostridium difficile Isolates without Requirement for Anaerobic Culture Conditions

2014 ◽  
Vol 52 (9) ◽  
pp. 3259-3263 ◽  
Author(s):  
Jennifer L. Cadnum ◽  
Kelly N. Hurless ◽  
Abhishek Deshpande ◽  
Michelle M. Nerandzic ◽  
Sirisha Kundrapu ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Culture Medium ◽  
Culture Conditions ◽  
Anaerobic Culture ◽  
Selective Culture Medium

Inhibition of proliferative growth in glioma cells by calmodulin antagonists

1986 ◽  
Vol 65 (1) ◽  
pp. 74-79 ◽  
Author(s):  
Nobuyuki Suzuki ◽  
Tetsuo Kanno ◽  
Yutaka Nagata ◽  
Taiji Kato
Keyword(s):  
Culture Medium ◽  
Deoxyribonucleic Acid ◽  
Control Level ◽  
Glioma Cells ◽  
Culture Conditions ◽  
Calmodulin Antagonists ◽  
High Concentration ◽  
Content Type ◽  
Chemically Induced ◽  
Proliferative Growth

✓ The effects of several calmodulin antagonists, such as N-(6-aminohexyl)-5-chloro-1-naphthalene-sulfonamide hydrochloride (W-7) and its dechlorinated structural analogue (W-5), on the growth and proliferation of cultured and transplanted glioma (GA-1, chemically induced from rat glioblasts) were evaluated. Under culture conditions, the concentration of W-7 necessary to exert 50% inhibition of GA-1 glioma cell growth was 50 µM. However, W-5, with a lower binding affinity to calmodulin than W-7, caused no definite inhibition of the proliferation of GA-1 cells in culture. When a low concentration of W-7 (12.5 µM) was added to the culture medium, deoxyribonucleic acid (DNA) synthesis in the GA-1 glioma cells was not markedly affected, whereas both ribonucleic acid (RNA) and protein syntheses were strongly suppressed on incubation for 24 hours. When a high concentration of W-7 (25.0 to 75.0 µM) was applied to the medium, synthesis of DNA, RNA, and protein was distinctly inhibited. When W-7 (50.0 µM) was added to the incubation medium, the calmodulin concentration in the cultured GA-1 was reduced to as much as half the control level within 2 hours, and thereafter remained at this level. Whereas control rats intraperitoneally transplanted with GA-1 cells could survive for 14 to 21 days, daily intraperitoneal injections of W-7 at concentrations of 1.0, 3.0, and 10.0 mg/kg body weight prolonged the survival span to between 21 and 26 days; this corresponded to an increased life span of about 40% compared to the controls.

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