杂色鲍原代细胞培养方法的优化及其在基因功能研究中的应用

优化了杂色鲍原代细胞的培养条件,并利用原代细胞进行了RNAi和外源基因表达。结果表明,较高的渗透压对血淋巴细胞培养起重要作用,淋巴细胞分离液分离血淋巴细胞更利于其生长。原代细胞培养至第5天,在细胞培养液中添加50μg My D88 dsRNA,能够显著降低My D88 mRNA的表达。p EGFP-N1转染培养5 d的鳃细胞,5 d后可以检测到绿色荧光。该研究表明可以成功培养杂色鲍血和鳃原代细胞,并且利用原代细胞可以进行功能基因表达量调低或调研究,为功能基因注释提供了便利条件。     

5种七彩神仙鱼5S rDNA序列的比较

采集鳍条,设计引物,提取了5种(黑格尔、盖子红、鸽子红、天子蓝、红点绿)七彩神仙鱼(Symphysodon)的5S r DNA,分析了其序列。结果表明:5种七彩神仙鱼均具有201 bp和401 bp 5S r DNA结构单元,而黑格尔(野生)、盖子红和鸽子红分别具有300,299,338 bp 5S r DNA结构单元。其编码区高度保守,而非转录间隔区差异显著,特别是黑格尔(野生)300 bp、盖子红299 bp和鸽子红338 bp的非转录间隔区。5种七彩神仙鱼G+C碱基的含量很高。红点绿、盖子红、鸽子红和天子蓝4个人工品种与野生品种黑格尔的亲缘关系分别为97.53%、90.12%、88.89%和85.19%,其中红点绿与黑格尔的亲缘关系最近。     

九孔鲍选择群体F1的选择反应与现实遗传力估计

以中国九孔鲍（Haliotis diversicolor aquatili）养殖群体与日本野生群体杂交子二代为选育基础群体,采用群体选育方法进行九孔鲍的选择育种研究,以估计该养殖群体的选择反应与现实遗传力。截断选择壳长最大的10％个体作为选择组亲本（SS）,从基础群体中随机抽取相同数量的个体作为对照组亲本（sc）,选择强度为1．755,结果表明,选择组与对照组在卵径、受精率方面无显著的差别（P〉0．05）,但选择组在幼体附着率、幼体变态率、幼体成活率及稚贝早期生长方面与对照组相比表现出不同程度的生长优势（P〈0．05）,选择组50日龄的稚鲍成活率达到35．07％,显著大于对照组23．62％（P〈0．05）,40—270日龄的选择组壳长选择反应和现实遗传力变化范围分别为13．64％～81．88％和0．21～0．53,平均为31．63％和0．36,选择组平均壳长显著大于对照组（P〈0．01）。研究初步显示,养殖群体的选择反应极显著,群体选育是养殖九孔鲍遗传改良的有效途径。     

Shell marking by artificial feeding of the tropical abalone Haliotis asinina Linne juveniles for sea ranching and stock enhancement

A method of marking abalone (Haliotis asinina Linne) for sea ranching and stock enhancement purposes was developed. Three‐month‐old abalone juveniles (11.8‐mm shell length, 0.28 g) were fed artificial diets for 1, 2, or 3 weeks. The width of the bluish‐green shell band produced by abalone juveniles was 1.7, 2.6, and 4.2 mm after 1, 2, or 3 weeks of feeding respectively. The growth and survival of juveniles fed artificial diets did not differ from that of juveniles fed the seaweed Gracilariopsis bailinae (control). Feeding the diet‐fed juveniles with the seaweed thereafter produced the natural brownish shell, thus forming a sandwiched bluish‐green band. An experimental release in outdoor tanks with natural growth of seaweeds and diatoms, and in a marine reserve showed that the shell band remained clear and distinct, indicating the usefulness of this shell marking method in sea ranching and stock enhancement of abalone.     

Physiological metabolism of Haliotis discus hannai Ino under different light qualities and cycles

The goal of this experiment was to examine the effects of different light qualities (red, white, blue) and cycles (12L:12D, 8L:16D, 4L:20D, 0L:24D, 16L:8D) on the survival, growth, metabolism and antioxidant defence system of Haliotis discus hannai Ino. Under red and white light at 4L:20D, the body weight‐specific growth rate of abalones was significantly higher than that of the 0L:24D group (P < 0.05). At 16L:8D, succinate dehydrogenase activity decreased and lactate dehydrogenase and alanine aminotransferase activity and the lactic acid content significantly rose (P < 0.05) in the red and white light groups. Under blue light, reactive oxygen species and malondialdehyde content for each light cycle were significantly higher than those of the red and white groups (P < 0.05). Under blue, white and red light, total antioxidant capacity (T‐AOC), superoxide dismutase (SOD) activity, glutathione peroxidase (GP_X) activity and reduced glutathione (GSH) content gradually rose with increased duration of light exposure. Under red light, significant differences in these parameters were detected between the 4L:20D and 16L:8D groups (P < 0.05). Under blue and white light at 16L:8D, values of T‐AOC, SOD, GP_X activity and GSH content were significantly lower than those of the 12L:12D group (P < 0.05). Overall, these results indicate that red light at light cycles of 4L:20D and 8L:16D is optimal for the culture and production of H. discus hannai, as these conditions promoted food intake, growth and health maintenance as well as normal physiological metabolism of the organism.     

Thermal preference and tolerance of green abalone Haliotis fulgens (Philippi, 1845) and pink abalone Haliotis corrugata (Gray, 1828)

The thermoregulatory behaviour of green abalone Haliotis fulgens and pink abalone H. corrugata was investigated. Haliotis fulgens juveniles ranging in wet weight from 3.0 to 3.3 g and from 28.7 to 30.5 mm shell length and of H. corrugata 2.0 g and 25.7 mm in shell length were exposed to 19°C for 30 days in a flow‐through water system. Temperature preference was determined in a horizontal thermal gradient and was found to be 25.4°C for green abalone and 25.0°C for pink abalone. Displacement velocity was 4.3 cm h⁻¹ for H. fulgens and 12.8 cm h⁻¹ for H. corrugata. The optimum temperature for growth calculated for both abalone species was 24.6 and 24.5°C respectively. The critical thermal maxima (CTMax) of H. fulgens and H. corrugata were determined as a measure of thermal tolerance. Abalones were subjected to increasing water temperatures at a rate of 1°C on 30 min until they detached from the substrate. The CTMax at 50% were 33.6 and 32.0°C for green and pink abalone respectively. The results are discussed in relation to site selection and commercial rearing.     

九孔鲍北方青苗及生长的探讨

探讨了九孔鲍(Haliotisdiversicolorsuertexta)在北方自然繁殖季节育苗的条件、发育时间和生长速度。2004年8月1日,用紫外线照射海水诱导产卵和人工授精的九孔鲍受精卵,经过60天培育,获6～12mm的幼鲍50万只。     

应用双抗夹心ELISA法检测皱纹盘鲍致病病原—创伤弧菌的研究

以皱纹盘鲍脓足病致病病原－ － 创伤弧菌为抗原,制备兔抗血清,抗体纯化后以辣根过氧化物酶标记,建立检测创伤弧菌的双抗夹心ＥＬＩＳＡ 检测法。结果表明,双抗夹心ＥＬＩＳＡ 法有较高的灵敏度,可检出含菌１０４ 个／ｍｌ 的菌悬液。与创伤弧菌对照菌株有明显的阳性反应不与副溶血弧菌、溶藻胶弧菌、河流弧菌等８ 株对照菌株产生影响检测结果的交叉反应。应用该法检测３０ 份病鲍样品,阳性检出率为６６ ．７ ％ 。     

Effect of ammonia on the growth rate and oxygen consumption of juvenile greenlip abalone, Haliotis laevigata Donovan

Juvenile greenlip abalone, Haliotis laevigata, (mean whole weight 4.48±1.9 g, mean±s.d., n=953) were highly sensitive to ammonia as indicated by depressed growth rate and food consumption measured over 2–3 months in bioassay tanks. For growth rate expressed on a whole weight basis, the EC₅ and EC₅₀ values (5 and 50% growth reductions) were 0.041 mg FAN l⁻¹ (Free Ammonia–Nitrogen) and 0.158 mg FAN l⁻¹, respectively. Shell growth rates declined over the entire experimental range (0.006–0.188 mg FAN l⁻¹). At the end of the bioassay, groups of abalone were transferred to respiratory chambers. Oxygen consumption rate increased to a maximum of 188% of control values at 0.235 mg FAN l⁻¹ and decreased slightly at the highest concentration of 0.418 mg FAN l⁻¹.     

杂色鲍的血细胞

通过光镜、电镜技术，比较了正常状态和受到细菌刺激后杂色鲍血细胞的变化。结果显示，正常状态下的杂色鲍血细胞分为三种类型：颗粒细胞、透明细胞和小细胞；杂色鲍血细胞具有很强的免疫功能，对外界刺激敏感；受到细菌刺激后杂色鲍的血细胞在结构和形态上均发生显著的变化。并推测，颗粒细胞的颗粒中含有大量的水解酶类和抗菌肽等免疫因子，发挥主要的免疫作用。