Development and application of a flexible controller in yeast fermentations using pO2 cascade control

The development and application of a flexible process controller in fed‐batch yeast fermentations using pO₂ cascade control was performed. A new algorithm for fed‐batch fermentations using pO₂ cascade control was developed, the concept of which could be used as a realizable solution in fermentation systems equipped according to the basic configuration. The algorithm is based on the combined influence of pO₂ and pH on the substrate feeding intensity. To test and develop this algorithm, Saccharomyces cerevisiae DY 7221 and Candida tropicalis CK‐4 fermentations were carried out. As a result of the use of the combined algorithm, the specific growth rate and productivity grew in both fermentations. In this case, the effect of the use of the algorithm was most pronounced in the C. tropicalis fermentation.     

Synthesis of the Human Insulin Gene: Protein Expression,Scaling Up and Bioactivity

Abstract

Optimized Synthetic human insulin gene was preferred to easy of cloning, plasmid stability, and protein expression away from the native sequence and its rare codons. Two steps to obtain the insulin, so we assembled the gene of 293 bp using a battery of overlapped synthetic oligos, then cloned into pET101directional TOPO expression vector downstream to the T7 promoter. The proinsulin products were produced as inclusion bodies in E. coli at a level of 10%. The batch cultivation of the strain yielded 6 g/L, while the high cell density of fed‐batch cultivation yielded 46 g/L. The proinsulin purification yielded 110 mg/gram cell weight, and 1.3 mg/gram of a bioactive insulin. The native insulin was generated by enzymatic conversion of chemically processed proinsulin. The produced insulin was matched with that of a commercial aqueous version at a level of enzyme immunoassys, SDS‐PAGE, RP‐HPLC, and bioactivity. The present results showed that the produced insulin has a comparable biochemical and potency similar to that of commercial one.     

Induction and inhibition of meiotic maturation of follicle-enclosed porcine oocytes in vitro

Isolated porcine Graafian follicles which were explanted in vitro and maintained in organ culture were used as a test-system for the meiosis-inducing action of PMSG and hCG. The addition of either PMSG or hCG alone (10 or 20 IU/ml, respectively) to the culture medium was not effective, whereas the simultaneous administration of these hormones ($\text{15}\text{15}\text{IU/ml}$) induced resumption of meiosis in 90.3% ($\text{37}\text{41}$). The same hormone concentrations were used in a second series of experiments in which the inhibition and induction of meiosis of isolated oocytes were tested by transferring them into host follicles. In host follicles containing up to 12 foreign eggs, which were cultured in control media, meiosis was prevented in 86.0% of all oocytes ($\text{104}\text{121}$). By adding PMSG (15 IU/ml) simultaneously with hCG (15 IU/ml) to the medium, meiosis was induced in 95.0% of all oocytes ($\text{133}\text{140}$).The assumption is made that the signal initiating resumption of meiosis of the isolated and transferred oocytes is mediated by the follicular fluid, since intimate contact with the membrana granulosa of the host follicle was prevented by using a roller technique.