马氏珠母贝IGF2BP1基因的鉴定及对矿化相关基因表达的影响

胰岛素样生长因子2结合蛋白( IGF2BPs)是一种重要的RNA代谢调控因子,涉及多个不同的生物学过程。本研究鉴定了一个马氏珠母贝IGF2BP1基因,命名为PfIGF2BP1-1,该基因cDNA全长为7348 bp,ORF长 1818 bp,编码605个氨基酸。多序列比对和系统进化分析表明,PfIGF2BP1-1有2个RRM结构域和4个KH结构域,与其他物种来源的IGF2BPs同源。实时荧光定量PCR结果显示,PfIGF2BP1-1 在马氏珠母贝的8个组织中均有表达,在肌肉组织表达最高,其次为足和外套膜组织。利用pET-32a 原核表达载体构建了含有PfIGF2BP1-1成熟多肽区的重组质粒并成功表达蛋白。在IPTG浓度为0.5 mmol/L,37 ℃培养8 h的条件下诱导表达PfIGF2BP1-1蛋白最佳。PfIGF2BP1-1蛋白刺激马氏珠母贝外套膜原代细胞,引起壳基质蛋白基因Accbp、MSI7、Nacrein的表达水平显著上升(p < 0.05),表明PfIGF2BP1-1蛋白可诱导壳矿化相关基因的表达参与马氏珠母贝的生物矿化过程。     

氮素形态对紫花苜蓿盛花期生长及结瘤固氮的影响

在完全营养液条件下,采用砂培法,研究了3种氮素形态配比NO3--N,NH4+-N和NO3--N∶NH4+-N为1∶1和0、105、210mg/L 3个氮素水平对紫花苜蓿盛花期生长及结瘤固氮的影响。结果表明:不同形态氮素处理下紫花苜蓿的株高、生物量、根瘤数、根瘤重、固氮酶活性和全氮含量均显著高于CK。同一氮素水平下,株高表现为NO3--N和NH4+-N混合培养下效果最好,NO3--N培养下次之,NH4+-N培养下最低;生物量、根瘤数、根瘤重、固氮酶活性和全氮含量NO3--N和NH4+-N混合培养下效果最好,NH4+-N培养下次之,NO3--N培养下最低;茎叶比则是NO3--N培养下最大,NH4+-N培养下次之,NO3--N和NH4+-N混合培养下的最低,各氮素形态处理间差异显著(P0.05)。随着氮素水平的增加,各形态配比下紫花苜蓿的株高、生物量、根瘤数、根瘤重、固氮酶活性和全氮含量呈现增加的变化趋势,茎叶比呈现减小的变化趋势。NO3--N+NH4+-N的浓度为210mg/L时,紫花苜蓿的株高、生物量、根瘤数、固氮酶活性和全氮含量取得最大值,茎叶比取得最小值,根瘤重则在NO3--N+NH4+-N的浓度为105mg/L时取得最大值,紫花苜蓿在混合态氮培养下生长最好。     

精子膜蛋白受精素(Fertilin)研究进展

精子膜蛋白受精素(Fertilin)是参与精卵质膜间相互作用最有特征性的候选分子之一。受精素由α、β两个亚基组成,为精子表面的异二聚体跨膜糖蛋白。fertilin-α(Fα)、Fertilin-β(Fβ)可能在精子发生、精子成熟、精子获能及精卵结合和融合中发挥重要作用。     

Cortisol does not affect hepatic α- and β-adrenoceptor properties in rainbow trout (Oncorhynchus mykiss)

Distribution and function of hepatic - and -adrenoceptors were examined in rainbow trout (Oncorhynchus mykiss) injected with slow release hydrogenated coconut oil implants alone (sham) or containing cortisol. - and -Adrenoceptors were assayed on purified hepatic membranes 10–14 days post-implantation using ³H-prazosin () and ³H-CGP (). At 10–14 days, plasma cortisol values were significantly elevated to approximately 220 compared with 35.0 ng ml^-1 in cortisol implanted vs. sham trout. No significant differences were found between any of the experimental groups for either the affinity (K_d) or maximal number of binding sites (B_max) for either receptor type. Epinephrine significantly stimulated glucose release from hepatocytes isolated from sham injected trout, but not from cortisol-treated fish. Epinephrine-induced glucose release was blocked by both - and -antagonists. These studies do not support the hypothesis that rainbow trout exposed to chronic cortisol alter properties of hepatic adrenoceptors.     

Renibacterium salmoninarum bar forms: characterization, occurrence, and evidence of a host response to a R. salmoninarum infection

Unique-staining Renibacterium salmoninarum (Rs) cells, termed bar forms, first observed in a coho salmon, Oncorhynchus kisutch (Walbaum), in 1983, could not be cultured, making their characterization difficult and significance obscure. They can be detected only by the fluorescent-antibody technique (FAT) and their numbers estimated only by a quantitative FAT (QFAT). Data collected over a 10-year period showed that bar forms were observed only in vivo and appeared associated with a host response. Bar forms were observed in 10 salmonid species from five countries and in fish from < 1 g to spawning adults. They were observed in 50.1% of kidney smears prepared from 10,061 Rs positive chinook, O. tshawytscha (Walbaum), coho, and Atlantic salmon, Salmo salar L. Bar forms were shown to be Rs cells based on absorption studies, their reaction with an Rs-specific FAT and enzyme-linked immunosorbent assay, and a transition from 'typical' Rs cells to bar forms in naturally and experimentally infected fish. Bar forms were determined to be non-virulent, damaged or dead Rs cells, based on fluorescence and electron microscopy observations, the inability to culture them, and mortality data. Bar forms appeared to represent visual markers of recovery from an Rs infection.     

棘头梅童鱼肝型脂肪酸结合蛋白基因克隆及组织表达分析

为进一步了解肝型脂肪酸结合蛋白基因(L-FABP基因)在棘头梅童鱼(Collichthys lucidus)营养调控、脂肪酸代谢和免疫应答等方面的功能作用,以棘头梅童鱼全基因组组装数据为基础,获得L-FABP基因的含开放阅读框的cDNA序列,利用生物信息学方法分析编码的蛋白质结构及进化关系,采用荧光定量(qRT-PCR)技术研究了不同组织中L-FABP基因的表达情况。结果表明,L-FABP基因cDNA序列为475 bp,其中5’非翻译区为51 bp,3’非翻译区为40 bp,开放阅读框为384 bp,编码127个氨基酸。其结构由10个β-折叠链和2个α-螺旋链组成,在氨基酸3~126位点间具有保守的脂质/胞质性脂肪酸结合结构域。棘头梅童鱼L-FABP基因与大黄鱼、条纹狼鲈同源性较近,与绿河鲀、尼罗罗非鱼和斑马鱼同源性较远。L-FABP基因在棘头梅童鱼肝脏、肠道、眼、大脑、心脏、肌肉、胃、鳃、脾脏、头肾和中肾中均有表达,其中肝脏中的L-FABP相对表达量最高,显著高于其他组织(P<0.05)。     

Binding and bioactivity of insulin in primary cultures of carp (Cyprinus carpio) hepatocytes

Isolated carp hepatocytes cultured in serum-free, chemically defined medium were used to investigate within the same cell preparation characteristics of the binding of insulin as well as effects of insulin on cellular metabolism. The binding of human [¹²⁵I]-insulin to carp hepatocytes was studied in kinetic, saturation and displacement experiments. A dependency of insulin binding on the collagenase used for cell isolation was demonstrated. Insulin binding decreased during the first 12h of culture but remained constant during the following 12h. The kinetic experiments revealed that [¹²⁵I]-insulin binding reached a steady state within 20–30 min of incubation. The mathematical analysis of the saturation experiments demonstrated the existence of two populations of binding sites, one with high affinity (K_d1 = 5.5 pM) and low capacity (B_max1 = 0.14 fmol/mg protein or 77 binding sites/cell) and one with low affinity (K_d2 = 2.4 nM) and high capacity (B_max2 = 17.6 fmol/mg protein or 9623 binding sites/cell). In competition experiments, 312 pM [¹²⁵I]-insulin was displaced by cold insulin, IGF-I and IGF-II with IC₅₀ values of 2.2, 7.9 and 20.3 nM, respectively. Glucagon was without effect. Binding of insulin to carp hepatocytes resulted in a significant reduction of glucose release and a significant increase of protein synthesis as of de novo fatty acid synthesis. dedicated to Prof. Dr. W. Hanke on the occasion of his 65^th birthday.     

Insulin and IGF-I receptors and tyrosine kinase activity in carp ovaries: changes with reproductive cycle

Insulin and insulin-like growth factor I (IGF-I) receptors from carp ovaries were semipurified with wheat germ agglutinin at different moments of the reproductive cycle and their binding characteristics and tyrosine kinase activity were studied. Specific receptors for insulin and IGF-I were found. IGF-I receptors presented higher binding (23.8 ± 1.5%), number of receptors (965 ± 20fm/mg) and affinity (K_D 0.24 ± 0.03nM) than those shown for insulin receptors (4.1 ± 1%, 530 ± 85fm/mg and 0.85 ± 0.1nM, respectively). Insulin and IGF-I receptors have a tyrosine kinase activity which is not different from that found in muscle of the same species. Seasonal changes were found in binding, with maximum values for insulin and IGF-I reached at the end of pre-spawning period (June). However, while IGF-I binding was observed in all stages, insulin binding decreased in autumn and disappeared in winter, which suggests a different role for the two peptides in ovarian physiology.     

Characterization of acetylcholinesterase and its molecular forms in organs of five freshwater teleosts

The activity and molecular forms of acetylcholinesterase (AChE, EC 3.1.1.7) were characterized in the brain, heart, white skeletal muscle and liver of the Siberian sturgeon (Acipenser baeri), European eel (Anguilla anguilla), wels (Silurus glanis), common carp (Cyprinus carpio) and silver carp (Hypophthalmichthys molitrix), belonging in four families. The brain had the highest (between 183.2±10.6 and 1361.2±189.7 mU mg^-1 protein and between 4.7±0.4 and 23.5±2.7 U g^-1 wet tissue) and the liver the lowest (between 25.3±1.9 and 126.5±22.7 mU mg^-1 protein and between 1.0±0.1 and 4.5±0.5 U g^-1 wet tissue) activity in all these fish, with the exception of the wels, where the heart contained the smallest amount of AChE (26.2±7.5 mU mg^-1 protein and 1.2±0.3 U g^-1 wet tissue). The highest tissue AChE activity was found in the carp and lowest in the sturgeon. The solubility properties and molecular forms of AChE in the four tissues were studied by extraction in high-salt medium (1.0 M NaCl) with and without the detergent Triton X-100 (0.5%, v/v). The proportions of detergent-soluble (DS) and salt-soluble (SS) forms of AChE varied considerably from one species to another, but a general tendency could be observed: the proportion of DS AChE was generally higher in the brain (between 70.4±4.1 and 82.5±1.2%) and lower in the heart (between 12.0±2.0 and 51.3±2.3%) and skeletal muscle (between 16.0±2.0 and 64.4±0.7%). Velocity sedimentation centrifugation revealed that most tissues contained the G₄ or A₄ form. Three tissues (brain, heart, and liver) of the Siberian sturgeon contained a relatively high proportion of the G1 form (between 24.3±2.4 and 28.6±4.9%), while the skeletal muscle of the wels contained only the most complex type of AChE, i.e., A₁₂. Amongst the tissues studied, the brain and liver mostly contained amphiphilic globular forms of the G₄ type, while the heart and skeletal muscle were rich in asymmetric forms (A₄ and A₁₂). It was concluded that the differences revealed in the activities of AChE and in the distribution of its molecular forms may be connected with the variation in behavioral habits (feeding and swimming patterns), and the developmental stage of the nervous system.     

两种形态蛹虫草菌株子实体及有效成分比较

在蛹虫草资源调查中,发现两种子实体形态迥异的蛹虫草菌株T和G。栽培试验表明,T菌株子实体矮化膨大,G菌株子实体为棒状．两者子实体长度、直径和产量差异显著。有效成分测定结果表明,菌株T和G子实体中粗多糖的含量分别为1．75％,1．77％,虫草酸为2．81％,2．57％,超氧化物歧化酶（SOD）酶活分别为30．43U／mgprotein、53．49U／mgprotein,过氧化物酶（POD）分别为31．26U／mgprotein,33．49U／mgprotein,过氧化氢酶（CAT）分别为126．35U／mgprotein、137．46U／mgprotein。其中,虫草酸含量、SOD酶活两者差异均显著。