草鱼(Ctenopharyngodon idellus)线粒体DNA控制区及其两侧tRNA基因的克隆与结构分析

动物线粒体ＤＮＡ控制区是线粒体基因组复制与基因表达的最主要的调控区．采用杂交和测序的方法对草鱼线粒体ＤＮＡ控制区进行定位、克隆并测定了控制区及其旁侧的ｔＲＮＡＰｈｅ、ｒＲＮＡＰｒｏ和ｒＲＮＡＴｈｒ三个基因的序列,与多种脊椎动物的相应序列进行了比较,并进行了结构分析．草鱼线粒体控制区全长９２７ｂｐ,含有与酵母和爪蟾线粒体启动子相似的序列,其ＣＳＢⅠ、ＣＳＢⅡ和ＣＳＢⅢ序列与其他几种动物的ＣＳＢ比较相当保守,ＴＡＳ与其回文基序可形成稳定的茎环结构,成为Ｈ－链复制的终止信号．草鱼线粒体ｔＲＮＡＰｈｅ、ｔＲＮＡＰｒｏ和ｔＲＮＡＴｈｒ可折叠成三叶草形二级结构,其基因具有许多不同于细胞质ｔＲＮＡ基因的结构特点,可能反映了线粒体ｔＲＮＡ与线粒体核糖体具有不寻常的作用方式     

siRNA-mediated knockdown of CiGRP78 gene expression leads cell susceptibility to heavy metal cytotoxicity

Heavy metal ion is one of the critical environmental pollutants accumulated in living organisms and causes toxic or carcinogenic effects once passed threshold levels. As an important member of Hsp70 (heat shock protein 70) family, the 78-kDa glucose-regulated protein (GRP78) can enhance cell survival rates remarkably under thermal stress. Recent studies also demonstrated that the expression of GRP78 enhances the cell survival under heavy metal stress. In this study, three most representative heavy metal ions, Pb^2 +, Hg^2 + and Cd^2 +, were used to stimulate Ctenopharyngodon idella kidney (CIK) cells. The results showed that cell viability under Pb^2 +, Hg^2 + and Cd^2 + stress decreased significantly. The longer and the greater the concentrations of stimulation from heavy metal ions, the higher the rate of cell death was observed. Among them, Hg^2 + is the most hazardous to cells. Under the same stress condition, Hg^2 + resulted in 50% of cell death, Cd^2 + (or Pb^2 +) led to 45% (or 35%) of cell death, respectively. Western immunoblotting indicated that C. idella GRP78 (CiGRP78) protein expression level was enhanced obviously in CIK cells under Pb^2 +, Hg^2 + and Cd^2 + stress, meaning CiGRP78 is involved in heavy metal cytotoxicity. To further study the role of CiGRP78 in cytoprotection, we designed the siRNA against CiGRP78 (from nucleotides + 788 to + 806) and transfected it into CIK cells to silence endogenous CiGRP78. The viability rate of CIK cells transfected with or without siRNA incubated with HgCl₂ for 12 h showed a significant decrease from 50% to 21%. Our results showed that CiGRP78 protects cells against heavy metal stimuli to some extent.     

Effect of melanin concentrating hormone on pigment and adrenal cells in vitro

Highly purified synthetic salmonid melanin concentrating hormone (MCH) and some analogs were investigated for their ability to concentrate the pigment in scale melanophores of the Chinese grass carp, Ctenopharyngodon idellus, to produce melanin dispersion in frog or lizard melanophores and to inhibit alpha-MSH in its action on mouse melanoma and rat adrenal glomerulosa cells in vitro. In the grass carp, MCH produced half-maximal pigment aggregation at 6 X 10(-11) M and its oxidized form at 7 X 10(-11) M. Replacement of the two methionines at position 3 and 6 with norvaline lowered the potency by a factor of 2.7 and with propargylglycine by a factor of about 7. Linear, Cys5,14-Acm-protected MCH was a full agonist of MCH but with a 345-fold lower potency. Iodinated MCH showed similar, low activity. In tetrapods, salmonid MCH and its analogs displayed only marginal pigment dispersion at concentrations greater than 10(-5) M. Alkali-treatment of MCH increased the pigment-dispersing potency by a factor of about 30 whereas the activity for pigment aggregation in the grass carp was destroyed. At high concentrations (10(-6), 10(-5) M) MCH also stimulated tyrosinase activity in B-16 mouse melanoma cells but did not modify the effects of alpha-MSH in this system. By contrast, when tested on rat adrenal glomerulosa cells, salmonid MCH had no effect alone but at a concentration of greater than 10(-10) M it slightly reduced corticosterone production by an alpha-MSH concentration of 10(-7) M. Aldosterone production was not affected and MCH did not influence the response to ACTH.     

Growth and energy budget in young grass carp, Ctenopharyngodon idella Val., fed plant and animal diets

Young grass carp held under laboratory conditions were fed either duckweed or tubificid worms. Food consumption ( C ), faecal production ( F ), excretion ( U ), metabolism ( R ) and growth ( G ) were estimated for fish fed both diets. Fish fed tubificid worms had higher growth rates than those fed duckweed, in terms of wet weight, dry matter, protein, lipid and energy. Fish fed duckweed consumed more wet and dry matter than did the fish fed tubificids, but rates of protein and lipid consumption were lower. The energy budget was estimated to be 100C = 9 F + 8 U + 61 R + 22 G for fish fed tubificids, and 100C = 35 F + 5 U + 51 R + 9 G for fish fed duckweed. Linear relationships existed between specific growth rate in wet weight and rate of food absorption (consumption minus faecal production) in dry matter, protein or energy. Food type did not have a significant effect on these relationships. It was concluded that differences in growth rate between grass carp fed different diets were mainly caused by differences in absorption rate.     

The bioenergetics of grass carp, Ctenopharyngodon idella (Val.): the influence of body weight, ration and dietary composition on nitrogenous excretion

Nitrogenous excretion by grass carp, Ctenopharyngodon idella (Val.), was measured in the form of ammonia and urea. Endogenous nitrogen excretion (ENE) was estimated as the daily rate of excretion by grass carp which had been starved for 2 days. ENE was scaled allometrically with body weight with weight exponents of 0.75 for ammonia, 0.63 for total nitrogen and 0.63 for the energy lost. The proportion of nitrogen attributable to urea was smaller than that attributable to ammonia and decreased from 25 to 12% as fish weight increased from 2 to over 10 g.
Linear relationships were found between daily rates of ammonia, total nitrogen and energy loss and daily rates of food intake. High carbohydrate and high lipid diets were not shown to have a protein-sparing action compared to a high protein diet. Differences in the amount of nitrogen excreted were explained by the differing nitrogen contents of the diets. Nitrogen budgets were erected and their implications discussed.     

Richness and diversity of helminth parasite communities in the common carp and in three more recently introduced carp species

It can be predicted that common carp Cyprinus carpio , as an introduced species, would harbour poor helminth infra-communities but that these would nevertheless be richer than those in three other species of carps introduced into Europe in recent decades. These predictions were tested by comparing the helminth infra-communities of common carp, Ctenopharyngodon idella, Hypophthalmichthys molitrix and Aristichthys nobilis reared under similar conditions in a fish farm. Data were presented on species richness of total helminth infra-communities, and on richness and diversity of gill, eye and intestinal infra-communities separately. These confirmed that common carp helminth infra-communities were poor and that they were no richer than those of the three other species of carp. This situation in the farm was considered to be representative of the situation in natural habitats in Europe. It is suggested that the poor infra-communities in common carp reflect the facts that their helminth communities are composed mainly of specialists, that they are not readily susceptible to invasion by specialists of other fish hosts and that, as introduced species, they have not had sufficient time to acquire many generalist helminths from native species of fish. The difference in time between the introduction of common carp into Europe centuries ago and the introduction of the other three species of carps decades ago is not sufficient to produce any significant differences in the richness and diversity of their helminth infra-communities.     

草鱼垂体STH细胞组织化学和超微结构研究

本文分别对草鱼性成熟前、后垂体ＳＴＨ细胞进行了组织化学和超微结构研究。垂体ＳＴＨ细胞多位于中腺垂体中部和背部,为嗜酸性细胞,用ＰＭＢ（ＰＡＳ－ＭＢ）和ＡＰＧ（ＡＢ－ＰＡＳ－ＯＧ）两种组织化学方法染色,对橙黄Ｇ阳性,对ＰＡＳ、ＡＢ阴性;电镜下电子密度较高,内质网绕核呈环形,分泌颗粒多而小。     

不同麻醉方法对降低草鱼捕捉应激的作用

正应激(Stress)指生物受到外源或内源刺激后内稳态改变而产生的一系列反应~[1]。在鱼类养殖中,各种操作对鱼产生的惊扰及环境变化均会导致鱼类应激~[2,3]。应激使鱼类代谢强度增加,免疫和生长性能下降~[4,5]。有关人类和哺乳动物的应激研究已积累了大量的资料,相比之下,鱼类应激方面     

草鱼与鲢鱼肥胖基因克隆与序列分析

肥胖基因产物leptin是调节哺乳动物摄食、能量代谢等生命活动的重要细胞因子。 应用RT-PCR和RACE法获得了草鱼(Ctenopharyngodon idellus)和鲢鱼(Hypophthalmichthys molitrix) leptin基因的全长cDNA序列分别为1 096 bp和1 176 bp,编码173和172个氨基酸。氨基酸序列同源性分析表明,草鱼和鲢鱼的leptin序列与其它鲤科鱼类leptin的同源性较高,而与其他鱼类的leptin同源性很低,但所有鱼类的leptin均含有用于形成二硫键的高度保守的半胱氨酸。系统进化树分析显示,草鱼和鲢鱼leptin与其他鱼类leptin聚于一进化分支。应用PCR和Genome Walker方法,进一步获得了草鱼和鲢鱼leptin基因的内含子和5′侧翼区序列。结果表明,获得的草鱼和鲢鱼leptin基因长度分别为2 129 bp和2 192 bp,含有与其他脊椎动物leptin相似的基因结构（含三个外显子和两个内含子）。本研究为深入研究鱼类肥胖基因结构功能关系与鱼类抗肥胖品系定向遗传选育奠定了良好的基础。