High-level expression ofβ-mannanase has been reported in Pichia pastoris under control of the GAP promoter.Two factors that strongly influence protein production and fermentation process development in Pichia pastori...High-level expression ofβ-mannanase has been reported in Pichia pastoris under control of the GAP promoter.Two factors that strongly influence protein production and fermentation process development in Pichia pastoris protein expression system are gene dosage and cultivation temperature.The aim of this research was to improve the expression level ofβ-mannanase in Pichia pastoris by proper increasing the gene dosage and decreasing the culture temperature.To this end,a panel of strains harboring different copy numbers ofβ-mannanase gene were obtained by multiple zeocin concentration gradients screening,the influence of gene copy number on the expression ofβ-mannanase in Pichia pastoris X33 was investigated.With the constitutive GAP promoter,the four copies strain exhibited a 4.04-fold higherβ-mannanase yield and a 1.83-fold higher total secretion proteins than the one copy strain,but an increase of the copy number above four resulted in a decrease of expression.Furthermore,the effects of culture temperature were studied in flask.The decreased culture temperature of four copies strain resulted in a 1.8-fold(26℃)and 3.5-fold(22℃)higherβ-mannanase activity compared to that at 30℃.A fed-batch strategy was successfully used for high cell-density fermentation andβ-mannanase activity reached 2124 U/mL after cultivation for 72 h in a 5 L fermenter.展开更多
This study was carried out at the vegetable research field, College of Agriculture, University of Duhok, during the growing season of 2009 using the eggplant local cultivar. And a factorial Randomized complete block d...This study was carried out at the vegetable research field, College of Agriculture, University of Duhok, during the growing season of 2009 using the eggplant local cultivar. And a factorial Randomized complete block design （F-RCBD）. The experiment consisted of two factors, the first was inoculations with and without humic acid, and the second factor was three levels of dry bread yeast （0.5, 10 gm/L）. The results revealed that the growth was significantly increased in term of plant height, branches number%, total chlorophyll and mineral content in leaves （N.P.K） and the yield in term of fruit number/plant, fruit weight （gm）, plant yield （gm/plant）, total yield （ton/hectare）, fruit length and fruit diameter. The interaction between humic acid and bread yeast was highly improved the growth and yield traits.展开更多
基金Project(31870115)supported by the National Natural Science Foundation of ChinaProject(2015JJ5006)supported by the Natural Science of Hunan Province&Changde City Joint Foundation,ChinaProjects(2015zzts268,ZY2015823)supported by the Fundamental Research Funds for the Central Universities,China
文摘High-level expression ofβ-mannanase has been reported in Pichia pastoris under control of the GAP promoter.Two factors that strongly influence protein production and fermentation process development in Pichia pastoris protein expression system are gene dosage and cultivation temperature.The aim of this research was to improve the expression level ofβ-mannanase in Pichia pastoris by proper increasing the gene dosage and decreasing the culture temperature.To this end,a panel of strains harboring different copy numbers ofβ-mannanase gene were obtained by multiple zeocin concentration gradients screening,the influence of gene copy number on the expression ofβ-mannanase in Pichia pastoris X33 was investigated.With the constitutive GAP promoter,the four copies strain exhibited a 4.04-fold higherβ-mannanase yield and a 1.83-fold higher total secretion proteins than the one copy strain,but an increase of the copy number above four resulted in a decrease of expression.Furthermore,the effects of culture temperature were studied in flask.The decreased culture temperature of four copies strain resulted in a 1.8-fold(26℃)and 3.5-fold(22℃)higherβ-mannanase activity compared to that at 30℃.A fed-batch strategy was successfully used for high cell-density fermentation andβ-mannanase activity reached 2124 U/mL after cultivation for 72 h in a 5 L fermenter.
文摘This study was carried out at the vegetable research field, College of Agriculture, University of Duhok, during the growing season of 2009 using the eggplant local cultivar. And a factorial Randomized complete block design （F-RCBD）. The experiment consisted of two factors, the first was inoculations with and without humic acid, and the second factor was three levels of dry bread yeast （0.5, 10 gm/L）. The results revealed that the growth was significantly increased in term of plant height, branches number%, total chlorophyll and mineral content in leaves （N.P.K） and the yield in term of fruit number/plant, fruit weight （gm）, plant yield （gm/plant）, total yield （ton/hectare）, fruit length and fruit diameter. The interaction between humic acid and bread yeast was highly improved the growth and yield traits.
