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51.
The large-scale production of recombinant biotherapeutics, particularly recombinant proteins, provides significant process and regulatory challenges to the biotechnology industry in order to meet the regulatory agencies stringent requirements in a cost-effective manner. Host cell derived nucleic acid causes problems from both a process and a regulatory perspective, as high molecular weight chromosomal DNA is responsible both for the viscosity of cell lysates, and it is a source of heterologous DNA sequences whose inclusion in the final product must be prevented. We have constructed a modified Escherichia coli JM107 expression host (JMN), containing a staphylococcal nuclease expression cassette, integrated into the host chromosome at the dif locus. The nuclease is expressed as a fusion to the ompA signal peptide, and is translocated to the periplasm of the cell, protecting the cytoplasmic nucleic acid from any toxic activity. The nuclease is released during cell lysis, where it subsequently acts to hydrolyse host nucleic acid present in the lysate. Results with this strain show that sufficient levels of nuclease activity are produced to completely auto-hydrolyse the host's chromosomal DNA to a size non-visible on 1% agarose gel, generating a markedly lower lysate viscosity. This provides a suitable methodology to remove heterologous DNA sequences early in the product stream and decrease lysate viscosity, improving the efficiency of downstream processing and product yield, whilst avoiding the addition of exogenous nuclease and its prohibitive costs at large-scale. 相似文献
52.
Angling-induced cardiac disturbance of free-swimming largemouth bass (Micropterus salmoides) monitored with heart rate telemetry 总被引:2,自引:0,他引:2
S. J. Cooke C. M. Bunt K. G. Ostrand D. P. Philipp D. H. Wahl 《Zeitschrift fur angewandte Ichthyologie》2004,20(1):28-36
The sub‐lethal effects of catch‐and‐release angling have been poorly studied because of the difficulties in monitoring physiological parameters in free‐swimming fish. Laboratory studies provide the opportunity to examine sub‐lethal effects in controlled environments, but do not incorporate site‐specific characteristics. In this study we angled free‐swimming largemouth bass (Micropterus salmoides) equipped with heart rate transmitters to exhaustion using rod and reel, and exposed fish to air for 30 s. Experiments were repeated at four water temperatures (13, 17, 21, and 25°C). These field data were compared with published findings from largemouth bass collected at the same water temperatures in a controlled laboratory setting using Doppler flow probes. Field collected heart rate data increased with increasing water temperatures (Q10 values 1.30–1.37). Pre‐disturbance heart rates were ~30% higher for free‐swimming fish in the field than previously collected laboratory data at the same water temperatures. Fish angled in the field exhausted ~40% more rapidly than fish chased in the laboratory. Maximal heart rate was ~15% higher for free‐swimming fish in the field than for data collected from laboratory restrained fish, but scope for heart rate was reduced by up to 20% in the field, especially at higher water temperatures. Heart rate in free‐swimming fish was highly variable at all times, obscuring clear recovery patterns. Conversely, laboratory cardiac parameters exhibited less variable patterns, peaking clearly following disturbances and recovering in about 135 min, independent of water temperature. Based upon these findings, we suggest that comprehensive studies incorporating both laboratory and field experiments are needed for truly understanding the effect of catch‐and‐release angling on fish. 相似文献
53.
W. T. Cooke C. H. J. Swan P. Asquith V. Melikian W. E. McFeely 《BMJ (Clinical research ed.)》1973,1(5849):324-327
Among 569 schoolchildren (386 boys and 183 girls) aged 14-17 years, 233 had serum alkaline phosphatase values of 30 K.A. units or greater. There was no significant difference in the results in Asian, white, or West Indian children. The mean values were significantly greater in boys than girls and both showed a fall in mean values with increasing age. Radiological rickets occurred in at least 4% of the survey, and was more common in Asians. Low calcium and high hydroxyproline excretion in most of those investigated and the response to vitamin D therapy suggests that most children with alkaline phosphatase levels above 30 K.A. units have rickets.Since the decline of the widespread supplementation of the diet with vitamin D, the demands of the physiological growth spurt for extra vitamin D in adolescents already on a borderline intake may be responsible for the great increase in “biochemical” rickets. Once the growth spurt is over the condition subsides but the results of impaired growth or permanent pelvic deformity will not necessarily be eradicated. 相似文献
54.
Chromate reduction by rabbit liver aldehyde oxidase 总被引:2,自引:0,他引:2
Chromate was reduced during the oxidation of 1-methylnicotinamide chloride by partially purified rabbit liver aldehyde oxidase. In addition to 1-methylnicotinamide, several other electron donor substrates for aldehyde oxidase were able to support the enzymatic chromate reduction. The reduction required the presence of both enzyme and the electron donor substrate. The rate of the chromate reduction was retarded by inhibitors of aldehyde oxidase but was not affected by substrates or inhibitors of xanthine oxidase. These results are consistent with the involvement of aldehyde oxidase in the reduction of chromate by rabbit liver cytosolic enzyme preparations. 相似文献
55.
K R Cooke 《Quarterly journal of experimental physiology and cognate medical sciences》1979,64(2):69-78
An oxygen electrode respirometer for determining the oxygen consumption of slices of mammalian renal cortex is described and assessed. Though rat renal cortical slices incubated in potassium-free medium for one hour lost 102 +/- 14 mmol of potassium/kg dry weight, there was only a small, nonsignificant fall in oxygen consumption. In contrast the oxygen consumption of slices incubated in potassium-free medium with 10 mmol.1-1 ouabain was markedly reduced (by 32 +/- 6%), while such slices lost 180 +/- 15 mmol of potassium/kg dry weight. These disproportionate effects on potassium loss and inhibition of oxygen consumption suggest that in renal cortical slices the loss of potassium in low potassium medium is not primarily due to inhibition of the conventional sodium pump. 相似文献
56.
We investigated the relationship between eggsize variation and (a) egg hatching success, (b) chick survival to fledging and
recruitment, and (c) adult female survival, over 12 years in the lesser snow goose (Anser caerulescens caerulescens). By comparing the means and variances of egg size for successful and unsuccessful eggs, our aim was to assess the relative
fitness of eggs of different sizes and to determine the type of selection operating on egg size in this species. As both egg
size and reproductive success vary with age in the lesser snow goose we controlled for the effects of female age. Egg-size
variation is very marked in this population, varying by up to 52% for eggs hatching successfully. However, there was no relationship
between egg size and post-hatching survival of goslings to fledging or recruitment, either within or between broods, pooling
across years. Egg size varied significantly between successful and unsuccessful clutches in only 2 of 33 individual year comparisons.
First-laid eggs surviving to onset of incubation, and eggs hatching successfully, were on average larger than unsuccessful
eggs, but this was probably due to the confounding effects of female age-specific and sequence-specific egg survival. Variance
of egg size differed significantly between successful and unsuccessful eggs in only 3 of 24, and 0 of 21, individual year
comparisons for pre- and post-hatching survival respectively. We therefore found little evidence for a relationship between
egg-size variation and offspring fitness, or for strong directional, normalising or diversifying selection operating on egg
size, in the lesser snow goose. In addition, there was only weak support for the hypothesis that egg-size variation is maintained
by temporal variation in selection pressure (sensu Ankney and Bisset 1973). It is likely that egg-size variation represents the pleiotropic expression of alleles affecting
more general physiological or metabolic processes. While this does not rule out the existence of alleles with more direct
effects on egg size we suggest that their contribution to heritable egg size is small. 相似文献
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