ApoB RNA编辑高效蛋白检测体系的构建

[目的]建立一个载脂蛋白B(apoB)RNA编辑蛋白检测体系。[方法]在慢病毒表达质粒载体pCSII-CMV-IRES-Neor中插入apoB RNA编辑保守序列,并在其2端嵌入红色荧光蛋白(DsRed)、绿色荧光蛋白(GFP)表达序列,以此慢病载体转染大鼠肝癌细胞系CBRH-7919获得稳定表达。采用共聚焦显微镜测序方法检测apoB RNA的编辑。[结果]目的指示基因能够在CBRH-7919细胞中稳定表达,表现出指征RNA编辑的多色特征。[结论]试验成功构建了在细胞水平上以颜色变化指示apoB RNA编辑的检测体系,该体系为快速检测以apoB RNA编辑为靶的降胆固醇药物筛选提供了基础。     

Three‐dimensional visualization of green fluorescence protein‐labelled Edwardsiella tarda in whole Medaka larvae

The invasive fish pathogen Edwardsiella tarda is common in aquatic environments and causes the environmentally and economically destructive emphysematous putrefactive disease called edwardsiellosis. In order to understand the organism's infection pathway, medaka larvae (Oryzias latipes) were immersion‐infected with E. tarda labelled with green fluorescence protein (GFP) and then visualized in three dimensions under confocal laser microscopy and light‐sheet fluorescence microscopy. Confocal microscopy revealed GFP‐labelled E. tarda in the mouth, head, gill bridges, gill cover, skin, membrane fin, gastrointestinal tract and air bladder, and in the caudal vein, somite veins, caudal artery and caudal capillaries. Light‐sheet microscopy additionally showed GFP‐labelled E. tarda in the pharyngeal cavity, muscle of the pectoral fin and cardiac atrium and ventricle. These findings suggest that during its infection of fish, E. tarda initially adheres to, and invades, the epithelial cells of the skin, gills and gastrointestinal tract (through the pharyngeal cavity); E. tarda then enters the blood vessels to access organs, including the air bladder and heart.     

Polymerisation of gluten proteins in developing wheat grain as affected by desiccation

The breadmaking quality of wheat is affected by the composition of gluten proteins and the polymerisation of subunits that are synthesised and accumulated in developing wheat grain. The biological mechanisms and time course of these events during grain development are documented, but not widely confirmed. Therefore, the aim of this study was to monitor the accumulation of gluten protein subunits and the size distribution of protein aggregates during grain development. The effect of desiccation on the polymerisation of gluten proteins and the functional properties of gluten were also studied. The results showed that the size of glutenin polymers remained consistently low until yellow ripeness (YR), while it increased during grain desiccation after YR. Hence, this polymerisation process was presumed to be initiated by desiccation. A similar polymerisation event was also observed when premature grains were dried artificially. The composition of gluten proteins, the ratios of glutenin to gliadin and high molecular weight-glutenin subunits to low molecular weight-glutenin subunits, in premature grain after artificial desiccation showed close association with the size of glutenin polymers in artificially dried grain. Functional properties of gluten in these samples were also associated with polymer size after artificial desiccation.     

Effect of dietary GroBiotic®‐A supplementation as a prebiotic on the intestinal microflora,growth performance,haemato‐serological parameters,survival rate and body composition in juvenile beluga (Huso huso Linnaeus, 1754)

The objectives of the present study were to investigate of GroBiotic^®‐A (GBA) on growth, autochthonous intestinal microbiota and haemato‐serological parameters of beluga juvenile. A total of 180 fish (40.82 ± 5.81 g) were fed diets containing graded levels of GBA (0, 5, 10 and 20 g kg^?1 diet) for 8 weeks. No significantly differences in body composition, total viable aerobic bacteria, mean cell volume, mean cell haemoglobin, mean cell haemoglobin concentration, neutrophils, eosinophils, monocytes, albumin, glucose, triglyceride, cholesterol, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase and lactate dehydrogenase were detected between treatment groups. The fish fed on 10 and 20 g kg^?1 GBA significantly showed higher lactic acid bacteria, final weight, body weight increases, weight gain and lower feed conversion ratio compared with the control and 5 g kg^?1 groups. The group fed 20 g kg^?1 GBA showed a highly significant difference in condition factor, survival rate, final length, total red and white blood cells, lymphocytes, haematocrit, haemoglobin, total protein, total immunoglobulin and lysozyme activity. The specific growth rate of the treatment groups was significantly elevated compared with the control groups. These results indicated that GBA at level 20 g kg^?1 improved growth, welfare and survival of beluga juvenile.     

Dietary protein requirement of juvenile obscure puffer,Takifugu obscurus

A 10‐week feeding experiment was conducted to determine the optimum dietary protein requirement of juvenile obscure puffer (Takifugu obscurus). Six isoenergetic (20 MJ kg^?1 gross energy) diets were formulated to contain graded levels of 34%, 38%, 42%, 46%, 50% or 54% crude protein (as dry matter basis). The results showed final body weight, weight gain and specific growth rate (SGR) increased significantly with increasing protein levels up to 42% and then decreased thereafer. Second‐order polynomial regression analysis (y = ?0.0024x² + 0.1788x ? 1.3196, R² = 0.9032) indicated a maximum SGR at protein level of 37%. Feed conversion ratio (FCR) decreased with increasing levels of dietary protein up to 42% and increased thereafter. Second‐order polynomial regression analysis (y = 0.0054x² ? 0.4351x + 10.391, R² = 0.753) indicated a minimum FCR at protein level of 40%. Protein efficiency ratio (PER) of fish fed the 34%, 38% and 42% diets was significantly higher than that of fish fed the 46%, 50% and 54% diets, and broken‐line analysis indicated PER tended to decrease when dietary protein level was higher than 40%. Generally, whole body lipid content, total cholesterol, low‐density lipoprotein cholesterol and triacylglycerol decreased with increasing levels of dietary protein. Fish fed the 42% protein diet showed the highest essential amino acids (histidine, isoleucine, leucine, lysine and threonine) and non‐essential amino acids (aspartic acid and glutamic acid) in muscle. Based on the second‐degree polynomial regression analysis of SGR and FCR and broken‐line analysis of PER, the optimal dietary protein level of obscure puffer is estimated to be between 37% and 40% (% as dry matter basis).     

Comparison of Conventional Washing Processing and pH Shift Processing on Gelation Characteristics of Bighead Carp (Aristichthys nobilis) Muscle Proteins

In this study, gelation characteristics of bighead carp (Aristichthys nobilis) protein isolates prepared by pH shift processing were investigated as compared with those of surimi prepared by conventional wash processing. Results showed that the gel from alkali-aided protein isolate exhibited a higher breaking force than that from conventional washed surimi (p < 0.05). However, the gel from conventional washed surimi had higher whiteness (p < 0.05) and lower expressible moisture (p < 0.05). The rheological study showed that protein isolates exhibited different storage modulus and loss tangent patterns from conventional washed surimi during heating. Acid-aided processing could lead to higher denaturation and dissociation for fish muscle myosin and a coarser gel network from the sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) pattern and the scanning electron microscopy (SEM). Overall, there is a potential to apply pH shift processing, especially alkali-aided processing, to extract more proteins from bighead carp muscle for the production of surimi-based food.     

Breeding for increased grain protein and micronutrient content in wheat: Ten years of the GPC-B1 gene

To provide food and nutrition security for a growing world population, continued improvements in the yield and nutritional quality of agricultural crops will be required. Wheat is an important source of calories, protein and micronutrients and is thus a priority to breed for improvements in these traits. The GRAIN PROTEIN CONTENT-B1 (GPC-B1) gene is a positive regulator of nutrient translocation which increases protein, iron and zinc concentration in the wheat grain. In the ten years since it was cloned, the impacts of GPC-B1 allelic variation on quality and yield traits have been extensively analyzed in diverse genetic backgrounds in field studies spanning forty environments and seven countries. In this review, we compile data from twenty-five studies to summarize the impact of GPC-B1 allelic variation on fifty different traits. Taken together, the results demonstrate that the functional copy of the GPC-B1 gene is associated with consistent positive effects on grain protein, Fe and Zn content with only marginally negative impacts on yield. We conclude that the GPC-B1 gene has the potential to increase nutritional and end use quality in a wide range of modern cultivars and environments and discuss the possibilities for its application in wheat breeding.     

Effects of the dietary protein to energy ratio on growth,feed utilization and body composition in Macrobrachium nipponense

Oriental river prawn (Macrobrachium nipponense) has been widely cultured in Asian countries. However, its nutritional studies are very limited. In the present 8‐week study, we investigated the effects of dietary protein to energy ratio (P/E ratio) on the growth, feed utilization and body composition in juvenile M. nipponense (initial weight 0.302 ±0.03 g). Two‐factor experiment was designed and nine semi‐purified diets were formulated to contain three lipid levels (20, 80 and 140 g kg^?1) and three protein levels (330, 380 and 430 g kg^?1), producing P/E ratios from 16.5 to 23.4 mg KJ^?1 protein. The results indicated that the growth, survival rate and protein efficiency were dose dependently improved by the increased dietary lipid, but not dietary protein content. Increased dietary lipid content and/or protein content increased lipid accumulation in whole body, hepatopancreas and muscle, but did not change the feed intake and hepatopancreas weight. In conclusion, our present study indicated that M. nipponense is a species with relatively high‐energy requirement. It could utilize dietary lipid content up to 140 g kg^?1, while the dietary protein with more than 330 g kg^?1 would not promote growth and protein efficiency. Taken together, 330 g kg^?1 dietary protein and 140 g kg^?1 dietary lipid level with P/E ratio 16.49 could be optimum for M. nipponense.     

Effect of high inclusion of rendered animal by‐product ingredients on growth,digestibility and economic performances in climbing perch Anabas testudineus

A 70‐day growth trail was conducted to investigate the effects of inclusion of high levels of meat and bone meal (MBM) and protein concentrate (PC) on growth, digestibility and economic performances of climbing perch, Anabas testudineus. Four isonitrogenous diets were formulated by lowering the level of dietary fishmeal protein at 0 (D₁, control), 70 (D₂), 85 (D₃) and 100% (D₄) with a mixture of MBM and PC protein (1:1). Triplicate groups of 300 fish (mean weight of 0.80 g) stocked in each 40 m² pond and fed the respective test diets. A digestibility trial was conducted after the growth trial in indoor glass aquarium. The result showed that growth parameters were significantly decreased (P < 0.05) with fishmeal replacement levels. However, significant differences were not found in feed conversion ratio and survival of fish. No difference was also found in protein efficiency ratio among D1, D2 and D3. Similar to growth parameters, total fish production was highest in D1, intermediate in D2 and D3; and lowest in D4. Apparent digestibility coefficients of dry matter, protein and lipid were highest (P < 0.05) in D1 and lowest in D4. The economic analysis revealed that the benefit cost ratio was ranked by D₃ (1.81), D₂ (1.71), D₁ (1.66) and D₄ (1.46) respectively. Upon considering the overall performances and unavailability of finite protein sources, it can be concluded that 70–85% fishmeal could be replaced with a mixture of MBM and PC (1:1) in practical diets for climbing perch.     

Choline supplementation increased total body lipid gain,while surplus methionine improved growth and amino acid retention in adult Atlantic salmon (Salmo salar)

Methionine–choline‐deficient (MCD) mammals are known to accumulate liver TAG probably due to phosphatidylcholine (PC) deficiency and thus assembly of VLDL and transport of lipids from liver to peripheral organs. To assess whether supplementation of choline could spare methionine and secure a healthy liver metabolism, by reducing the endogenous PC synthesis without interfering with lipid transport and distribution, Atlantic salmon with initial BW of 700 g were fed adequate (1.9 g Met/16 gN) or surplus methionine (2.5 g Met/16 gN) diets of which were supplemented with choline or not for a period of 19 weeks. Fish fed the low‐methionine diets had reduced growth (p = .013) due to reduced protein gain (p = .007), while lipid gain slightly improved in fish fed the choline‐supplemented diets (p = .047). Also, feed conversion improved when fed surplus methionine (p < .001), while choline supplementation had no impact on feed conversion. No interaction between choline and methionine on growth performance or retention existed. Phospholipid status in liver and muscle was not affected by treatments, and no liver TAG accumulation occurred at the methionine levels used. Gene expression of ApoB100 necessary for assembling VLDL or pemt necessary for endogenous PC synthesis was unaffected by treatments. Capacity of methylation (MAT, BHMT) within the liver was not affected by treatment nor was the gene expression of enzymes in liver transsulfuration (CBS or CDO). Methionine status within liver was unaffected by treatments, while free methionine reduced in those fish fed the low‐methionine diets in muscle and plasma. Cystathionine and taurine were elevated when fed surplus methionine. Choline supplementation had no impact on sulphur amino acid metabolites in either tissue. Neither did choline supplementation improve TAG mobilization from liver to muscle. To conclude, choline does not improve endogenous phospholipid synthesis or transport of TAG from liver to muscle depot when added to diets containing 1.9 g Met/16 gN, while surplus methionine improved growth and protein retention, indicating that 1.9 g Met/16 gN is enough to support a healthy liver metabolism, but too low to support muscle protein deposition in adult salmon fed high plant protein diets for longer periods of time.