Altering the ribosomal subunit ratio in yeast maximizes recombinant protein yield

Background The production of high yields of recombinant proteins is an enduring bottleneck in the post-genomic sciences that has yet to be addressed in a truly rational manner. Typically eukaryotic protein production experiments have relied on varying expression construct cassettes such as promoters and tags, or culture process parameters such as pH, temperature and aeration to enhance yields. These approaches require repeated rounds of trial-and-error optimization and cannot provide a mechanistic insight into the biology of recombinant protein production. We published an early transcriptome analysis that identified genes implicated in successful membrane protein production experiments in yeast. While there has been a subsequent explosion in such analyses in a range of production organisms, no one has yet exploited the genes identified. The aim of this study was to use the results of our previous comparative transcriptome analysis to engineer improved yeast strains and thereby gain an understanding of the mechanisms involved in high-yielding protein production hosts. Results We show that tuning BMS1 transcript levels in a doxycycline-dependent manner resulted in optimized yields of functional membrane and soluble protein targets. Online flow microcalorimetry demonstrated that there had been a substantial metabolic change to cells cultured under high-yielding conditions, and in particular that high yielding cells were more metabolically efficient. Polysome profiling showed that the key molecular event contributing to this metabolically efficient, high-yielding phenotype is a perturbation of the ratio of 60S to 40S ribosomal subunits from approximately 1:1 to 2:1, and correspondingly of 25S:18S ratios from 2:1 to 3:1. This result is consistent with the role of the gene product of BMS1 in ribosome biogenesis. Conclusion This work demonstrates the power of a rational approach to recombinant protein production by using the results of transcriptome analysis to engineer improved strains, thereby revealing the underlying biological events involved.

Phase shift keyed systems based on a gain switched laser transmitter

Return-to-Zero (RZ) and Non-Return-to-Zero (NRZ) Differential Phase Shift Keyed (DPSK) systems require cheap and optimal transmitters for widespread implementation. The authors report on a gain switched Discrete Mode (DM) laser that can be employed as a cost efficient transmitter in a 10.7 Gb/s RZ DPSK system and compare its performance to that of a gain switched Distributed Feed-Back (DFB) laser. Experimental results show that the gain switched DM laser readily provides error free performance and a receiver sensitivity of -33.1 dBm in the 10.7 Gbit/s RZ DPSK system. The standard DFB laser on the other hand displays an error floor at 10(-1) in the same RZ DPSK system. The difference in performance, between the two types of gain switched transmitters, is analysed by investigating their linewidths. We also demonstrate, for the first time, the generation of a highly coherent gain switched pulse train which displays a spectral comb of approximately 13 sidebands spaced by the 10.7 GHz modulation frequency. The filtered side-bands are then employed as narrow linewidth Continuous Wave (CW) sources in a 10.7 Gb/s NRZ DPSK system.

Full-field feed-forward equalizer with adaptive system optimization

We numerically investigate the combination of full-field detection and feed-forward equalizer (FFE) for adaptive chromatic dispersion compensation up to 2160 km in a 10 Gbit/s on-off keyed optical transmission system. The technique, with respect to earlier reports, incorporates several important implementation modules, including the algorithm for adaptive equalization of the gain imbalance between the two receiver chains, compensation of phase misalignment of the asymmetric Mach-Zehnder interferometer, and simplified implementation of field calculation. We also show that in addition to enabling fast adaptation and simplification of field calculation, full-field FFE exhibits enhanced tolerance to the sampling phase misalignment and reduced sampling rate when compared to the full-field implementation using a dispersive transmission line.

Self-seeded gain-switched operation of an InGaN MQW laser diode

Self-seeded, gain-switched operation of an InGaN multi-quantum-well laser diode has been demonstrated for the first time. An external cavity comprising Littrow geometry was implemented for spectral control of pulsed operation. The feedback was optimized by adjusting the external cavity length and the driving frequency of the laser. The generated pulses had a peak power in excess of 400mW, a pulse duration of 60ps, a spectral linewidth of 0.14nm and maximum side band suppression ratio of 20dB. It was tunable within the range of 3.6nm centered at a wavelength of 403nm.

Noise and gain optimisation in bi-directionally pumped dispersion compensating amplifier modules

We perform optimisation of bi-directionally pumped dispersion compensating Raman amplifier modules. Optimal forward and backward pump powers for basic configurations using different commercially available fibers are presented for both single- and multi-channel systems. Optical signal-to-noise ratio improvement of up to 8 dB is achieved as a result of optimisation. © 2003 Published by Elsevier B.V.

Optimization of gain and SNR in bi-directionally pumped dispersion compensating amplifier modules

In this paper, we present an analysis and optimisation of the performance of bi-directionally pumped dispersion compensation modules acting as simultaneous Raman amplifiers, with optimal configurations for operation with different fibers commercially available. The ratio between forward and backward pump powers for minimum noise influence is obtained in each case, with improvements in the SNR of up to 8 dB when compared to a purely backward-pumped case.

Solving Ratio-Dependent Predator-Prey System with Constant Effort Harvesting using Variational Iteration Method

Due to wide range of interest in use of bio-economic models to gain insight into the scientific management of renewable resources like fisheries and forestry,variational iteration method (VIM) is employed to approximate the solution of the ratio-dependent predator-prey system with constant effort prey harvesting.The results are compared with the results obtained by Adomian decomposition method and reveal that VIM is very effective and convenient for solving nonlinear differential equations.

Full-field feed-forward equalizer with adaptive system optimization

We numerically investigate the combination of full-field detection and feed-forward equalizer (FFE) for adaptive chromatic dispersion compensation up to 2160 km in a 10 Gbit/s on-off keyed optical transmission system. The technique, with respect to earlier reports, incorporates several important implementation modules, including the algorithm for adaptive equalization of the gain imbalance between the two receiver chains, compensation of phase misalignment of the asymmetric Mach-Zehnder interferometer, and simplified implementation of field calculation. We also show that in addition to enabling fast adaptation and simplification of field calculation, full-field FFE exhibits enhanced tolerance to the sampling phase misalignment and reduced sampling rate when compared to the full-field implementation using a dispersive transmission line.

Design of a modular, high step-up ratio DC–DC converter for HVDC applications integrating offshore wind power

High-power and high-voltage gain dc-dc converters are key to high-voltage direct current (HVDC) power transmission for offshore wind power. This paper presents an isolated ultra-high step-up dc-dc converter in matrix transformer configuration. A flyback-forward converter is adopted as the power cell and the secondary side matrix connection is introduced to increase the power level and to improve fault tolerance. Because of the modular structure of the converter, the stress on the switching devices is decreased and so is the transformer size. The proposed topology can be operated in column interleaved modes, row interleaved modes, and hybrid working modes in order to deal with the varying energy from the wind farm. Furthermore, fault-tolerant operation is also realized in several fault scenarios. A 400-W dc-dc converter with four cells is developed and experimentally tested to validate the proposed technique, which can be applied to high-power high-voltage dc power transmission.

A novel 3-D segmentation algorithm for anatomic liver and tumor volume calculations for liver cancer treatment planning

Three-Dimensional (3-D) imaging is vital in computer-assisted surgical planning including minimal invasive surgery, targeted drug delivery, and tumor resection. Selective Internal Radiation Therapy (SIRT) is a liver directed radiation therapy for the treatment of liver cancer. Accurate calculation of anatomical liver and tumor volumes are essential for the determination of the tumor to normal liver ratio and for the calculation of the dose of Y-90 microspheres that will result in high concentration of the radiation in the tumor region as compared to nearby healthy tissue. Present manual techniques for segmentation of the liver from Computed Tomography (CT) tend to be tedious and greatly dependent on the skill of the technician/doctor performing the task. ^ This dissertation presents the development and implementation of a fully integrated algorithm for 3-D liver and tumor segmentation from tri-phase CT that yield highly accurate estimations of the respective volumes of the liver and tumor(s). The algorithm as designed requires minimal human intervention without compromising the accuracy of the segmentation results. Embedded within this algorithm is an effective method for extracting blood vessels that feed the tumor(s) in order to plan effectively the appropriate treatment. ^ Segmentation of the liver led to an accuracy in excess of 95% in estimating liver volumes in 20 datasets in comparison to the manual gold standard volumes. In a similar comparison, tumor segmentation exhibited an accuracy of 86% in estimating tumor(s) volume(s). Qualitative results of the blood vessel segmentation algorithm demonstrated the effectiveness of the algorithm in extracting and rendering the vasculature structure of the liver. Results of the parallel computing process, using a single workstation, showed a 78% gain. Also, statistical analysis carried out to determine if the manual initialization has any impact on the accuracy showed user initialization independence in the results. ^ The dissertation thus provides a complete 3-D solution towards liver cancer treatment planning with the opportunity to extract, visualize and quantify the needed statistics for liver cancer treatment. Since SIRT requires highly accurate calculation of the liver and tumor volumes, this new method provides an effective and computationally efficient process required of such challenging clinical requirements.^

Níveis de cálcio e relação cálcio: fósforo em rações para poedeiras leves de 24 a 40 semanas de idade

v. 17, n. 2, p. 206-216, abr./jun. 2016.

Evaluation of processed groundnut (Arachis hypogaea) cake meals in the feeding of Clariid catfish, Clarias gariepinus, fingerlings

Groundnut cake (GNC) meal is an important source of dietary protein for domestic animals with a cost advantage over the conventional animal protein sources used in aquaculture feed production. It would be useful to evaluate the effects of GNC processing methods on the density and nutritional values of processed GNC meals. The use of processed GNC meals in the diets of Clarias gariepinus fingerlings was evaluated. Seven iso-proteic and iso-caloric diets were formulated, replacing fish meal with roasted and boiled GNC meals, each at three inclusion levels of 30%, 35%, and 40%. Diet I is 100% fishmeal, Diet II is 30% roasted GNC meal, Diet III is 35% roasted GNC meal, Diet IV is 40% roasted GNC meal, Diet V is 30% boiled GNC meal, Diet VI is 35% boiled GNC meal and Diet VII is 40% boiled GNC meal. Results showed that the crude protein content of GNC meals was 40.5% and 40.8% in boiled and roasted GNC meals respectively; the lower protein content for processed GNC meals might be due to heat denaturation of the seed protein, with boiled GNC meal being more adversely affected. The mean weight gain of fingerlings fed roasted GNC meals ranged between 5.29 – 5.64 while for boiled GNC meals, it was between 4.60 – 5.22. Generally, fish performed better when fed diets containing roasted GNC meals, than boiled GNC meals, and compared favorably with fish fed fish meal based diet. Body mass increase, total feed increase, protein efficiency ratio and specific growth rate by C. gariepinus fingerlings in all diets, showed no significant differences, suggesting that processed GNC meals could partially replace diets for C. gariepinus fingerlings without adverse consequences. This study showed that processed GNC meals could partially replace fish meal up to 30% without significantly influencing fingerling growth and health. It is recommended that the use of fish meal as the main basal ingredient for fingerlings could be discontinued, since GNC meal was a cheaper alternative, and could replace fish meal up to 35%, without any significant adverse effects on the fingerling performance. KEYWORDS: Clarias gariepinus, Fingerlings, Groundnut cake meal, Nutrient utilization, Performance.

Production responses of reproducing ewes to a by-product-based diet inoculated with the probiotic Bacillus amyloliquefaciens strain H57

The potential application of the spore-forming probiotic Bacillus amyloliquefaciens strain H57 (H57) as a novel probiotic for ruminants was evaluated in reproducing ewes. Performance responses were determined by delivering H57 in a pelleted diet based mainly on palm kernel meal (PKM) and sorghum grain. PKM is an agro-industrial by-product with a reputation for poor palatability and the availability of the starch in sorghum grain can be limited in ruminants. The hypothesis was that H57 improves the feeding value of a relatively low quality concentrate diet. Twenty-four first-parity white Dorper ewes were fed PKM-based pellets manufactured with or without H57 (109 cfu/kg pellet) in late pregnancy. During this phase of late pregnancy, the H57 ewes ate 17% more dry matter (1019 vs 874 g/day, P = 0.03), gained more weight (194 vs 30 g/day, P = 0.008) and retained more nitrogen (6.13 vs 3.34 g/day, P = 0.01), but produced lambs with a similar birthweight (4.1 vs 4.2 kg, P = 0.73). Rumen fluid collected from H57 ewes in late pregnancy had higher pH (7.1 vs 6.8, P = 0.07), acetate : propionate ratio (3.4 vs 2.7, P = 0.04), lower ammonia (69 vs 147 mmol/L, P = 0.001) and total volatile fatty acid concentrations (40 vs 61 mg/L, P = 0.02). The digestibility of dry matter, organic matter and fibre were similar between the two groups. The lambs of the H57 ewes grew faster than those of the Control ewes for the first 21 days of lactation (349 vs 272 g/day, P = 0.03), but not thereafter. H57 can improve feed intake and maternal liveweight gain in late pregnancy of first-parity ewes fed a diet based on PKM.

Near complete genome sequence of the animal feed probiotic, Bacillus amyloliquefaciens H57

Bacillus amyloliquefaciens H57 is a bacterium isolated from lucerne for its ability to prevent feed spoilage. Further interest developed when ruminants fed with H57-inoculated hay showed increased weight gain and nitrogen retention relative to controls, suggesting a probiotic effect. The near complete genome of H57 is ~3.96 Mb comprising 16 contigs. Within the genome there are 3,836 protein coding genes, an estimated sixteen rRNA genes and 69 tRNA genes. H57 has the potential to synthesise four different lipopeptides and four polyketide compounds, which are known antimicrobials. This antimicrobial capacity may facilitate the observed probiotic effect.

Growth, feed intake, survival, and histological response of white shrimp Litopenaeus vannamei fed diets containing grains naturally contaminated with aflatoxin = Crecimiento, consumo de alimento, supervivencia y respuesta histológica del camarón blanco Litopenaeus vannamei alimentado con dietas de granos contaminados naturalmente con aflatoxinas

Two feeding trials were carried out to evaluate the effect of diets containing corn or peanut grains naturally contaminated with aflatoxins on the growth, feed intake, survival, and histological response of the white shrimp Litopenaeus vannamei. In trial 1, four experimental diets were formulated to contain 0, 500, 1000, and 2000 g kg–1 of total aflatoxins (TA) and fed to L. vannamei juveniles for 28 days. In trial 2, six experimental diets were formulated to contain 0, 10, 20, 40, 60, and 120 g kg–1 TA and fed to L. vannamei juveniles for 64 days. Feed intake and weight gain were significantly affected by the presence of aflatoxins from naturally contaminated grains. Feed conversion rate increased significantly from a level of inclusion of 60 g kg–1. Survival was significantly reduced only for shrimp fed diets supplemented with 1000 and 2000 g kg–1 TA. Shrimp exposed to higher aflatoxin inclusion levels presented significantly lower lipid vacuole levels in R-cells (12–28%), lower B-cell activity, and lower mitotic E-cell activity. Tubular epithelial atrophy increased from the inclusion level of 20 g kg–1. Hepatopancreatocyte sloughing was significantly higher in shrimp fed diets supplemented with 1000 and 2000 g kg–1 TA. It is worth noting that shrimp fed 40 g kg–1 TA presented a high hepatopancreatocyte sloughing coefficient. Based on these results we conclude that the presence of aflatoxins, even at low levels, reduces feed intake and weight gain, and alters the cells of the hepatopancreas. RESUMEN. Se llevaron a cabo dos bioensayos para evaluar el efecto de granos de maíz y maní contaminados naturalmente con aflatoxinas sobre el crecimiento, consumo de alimento, supervivencia y daños histológicos de juveniles del camarón blanco Litopenaeus vannamei. Para el bioensayo 1, se formularon cuatro dietas con 0, 500, 1000 y 2000 g kg–1 de aflatoxinas totales (AT) y se proporcionaron a juveniles de L. vannamei durante 28 días. Para el bioensayo 2, se formularon seis dietas con 0, 10, 20, 40, 60 y 120 g kg–1 AT y se proporcionaron a juveniles de L. vannamei durante 64 días. El consumo de alimento fue significativamente afectado por la presencia de aflatoxinas. La tasa de conversión alimenticia incrementó significativamente a partir de un nivel de inclusión de 60 g kg–1. La supervivencia fue significativamente reducida solamente en los camarones que fueron alimentados con las dietas suplementadas con 1000 y 2000 g kg–1 AT. Los camarones expuestos a los niveles de inclusión altos presentaron un menor nivel de vacuolas lipídicas en las células R (12–28%), y una menor actividad de las células B y de la actividad mitótica de las células E. La atrofia de los túbulos del epitelio se incrementó a partir de un nivel de inclusión de 20 g kg–1. La descamación de las células del hepatopáncreas fue significativamente mayor en camarones alimentados con las dietas suplementadas con 1000 y 2000 g kg–1 AT, mientras que para las dosis bajas no se observaron diferencias significativas, aunque en camarones alimentados a partir de 40 g kg–1 AT se observa un coeficiente de descamación alto. Con base en los resultados, se concluye que la presencia de aflatoxinas, incluso a niveles bajos, reduce el consumo de alimento y el aumento de peso y altera las células del hepatopáncreas.