紫外辐射灭活大黄鱼和黄姑鱼精子及其激活大黄鱼卵子发育为胚胎的效果

为探讨大黄鱼(Larimichthys crocea)和黄姑鱼(Nibea albiflora)精子的紫外辐射灭活适宜剂量及其激活大黄鱼卵子发育为胚胎的效果,以Ringer氏液为稀释液,按1:30稀释大黄鱼和黄姑鱼精子,采用自制紫外灭活装置[紫外辐射强度2200μW/(cm~2·s),紫外波长254 nm]对这2种鱼的精子进行紫外照射处理及活力测定,然后与正常大黄鱼卵子进行人工授精,授精后一部分卵未作冷休克处理,另一部分卵进行了冷休克处理(受精2 min 30 s,3℃海水,冷休克10 min),并进行了早期胚胎成活率和仔鱼孵化率的测定与比较。结果显示:1)大黄鱼和黄姑鱼精子的激活率与紫外照射处理时间呈负相关,精子的快速运动时间变化呈典型的Hertwig效应。2)未冷休克组中大黄鱼和黄姑鱼诱导的早期胚胎成活率与精子的紫外照射时间总体呈负相关,而仔鱼孵化率呈Hertwig效应。3)冷休克组中大黄鱼和黄姑鱼精子诱导的早期胚胎成活率和仔鱼孵化率随紫外照射时间的增加呈Hertwig效应,分别于2 min 20 s和1min 30 s时达相对峰值,此时,大黄鱼早期胚胎成活率和仔鱼孵化率分别为(38.3±4.3)%和(66.5±5.1)%,黄姑鱼早期胚胎成活率和仔鱼孵化率分别为(43.3±3.3)%和(67.7±6.3)%。分析认为,大黄鱼和黄姑鱼精子遗传失活的紫外辐射剂量分别以308 m J/cm~2和198 m J/cm~2为佳。本研究旨在为大黄鱼雌核发育技术的改进提供依据。     

Effect of dietary vitamin C on growth performance,body composition and biochemical parameters of juvenile Chu's croaker (Nibea coibor)

Vitamin C is an essential micronutrient for normal physiological and immune functions of fish. However, its requirements and effects in Chu's croaker (Nibea coibor) are currently unknown. A 56‐day feeding trial was conducted to evaluate the optimal dietary vitamin C requirements based on its effects on growth performance, body composition and biochemical parameters in juvenile Chu's croaker (14.17 ± 0.1 g). Six isoproteic (450 g/kg crude protein) and isolipidic (100 g/kg crude lipid) diets were formulated to contain 2.24 (basal diet), 39.03, 85.01, 171.16, 356.49 and 715.46 mg/kg of vitamin C. The results showed that fish fed on 171.16 mg/kg vitamin C diet had the highest growth performance and feed utilization. Fish fed on the basal diet had higher malondialdehyde (MDA) content and lower activities of antioxidant enzymes in the serum and liver as compared with those fed on vitamin C diets. Polynomial analysis indicated that the optimal dietary vitamin C requirements of juvenile Chu's croaker were 102.28, 98.21, 150.26, 165.38, 71.46, 176.19, 84.84 and 103.78 mg/kg based on weight gain, specific growth rate, liver storage, muscle storage, liver MDA content, liver alanine aminotransferase activity, liver alkaline phosphatase activity and liver superoxide dismutase activity, respectively. We recommend an inclusion level in the range of 71.46–150.26 mg/kg vitamin C in the diets of juvenile Chu's croaker for optimum growth performance, liver function, antioxidant capacity and innate immunity functions.     

东海黄姑鱼年龄与生长的初步研究

根据2004年2~4月在舟山沈家门市场采集的117ind黄姑鱼样品,对黄姑鱼的年龄与生长进行了初步研究。结果表明:①目前舟山渔场渔获物中黄姑鱼年龄组成主要以1~2龄的低龄组为主,主要体长在161~263mm之间,主要体重在175.3~354.8g之间;②黄姑鱼体长与体重的关系为:W=1.633×10-5L3.0357;③体长和体重的生长方程分别为:Lt=529.5[1-e-0.262(t-0.281)]和Wt=2597.4[1-e-0.262(t-0.281)]2.4164;④体长生长速度方程为:dL/dt=138.729e-0.262(t-0.281),体重生长速度方程为:dW/dt=1644.4056e-0.262(t-0.281)×[1-e-0.262(t-0.281)]1.4164,生长加速度方程分别为:d2L/dt2=-36.3470e-0.262(t-0.281)和d2W/dt2=430.8343e-0.262(t-0.281)×[1-e-0.262(t-0.281)]0.4164×[2.4164e-0.262(t-0.281)-1]。     

温度和盐度对日本黄姑鱼胚胎发育的影响

为了确定日本黄姑鱼（Nibea japonica）胚胎孵化的最适水温和盐度,采用不同水温和盐度对日本黄姑鱼受精卵进行了孵化试验。结果显示,在盐度为30的条件下,日本黄姑鱼受精卵在14～28℃范围内均能正常孵化,最适孵化水温为18～20℃。孵化时数与温度呈幂函数相关,孵化时间随温度升高而缩短。盐度试验表明,在水温为18℃的条件下,日本黄姑鱼受精卵在盐度25—32的海水中呈悬浮或者半悬浮状态,在盐度14～45范围内均能孵化,适宜孵化盐度为26～32。盐度对初孵仔鱼全长有显著影响,盐度较低组仔鱼全长显著大于高盐度组（P〈0．05）。     

浅色黄姑鱼脑垂体cDNA文库的构建

以浅色黄姑鱼脑垂体为材料,应用SMART~(TM) cDNA library construction技术,构建以真核表达载体pcD- NA3．1( )为基础的浅色黄姑鱼cDNA文库。利用含Sfi I B酶切位点的oligo(dT)引物合成cDNA第一链,利用含Sfi I A酶切位点的SMART核苷酸作为cDNA第一链在mRNA5′端延伸出去的模板,采用LD PCR引物合成双链cDNA,双链cDNA用Sfi I酶切和过柱分级分离后,与引入Sfi I酶切位点的pcDNA3．1( )-Sfi I载体进行连接,转化大肠杆菌TOP10感受态细胞,构建成脑垂体全长cDNA文库。经过质量鉴定,得到的原始cDNA文库含有约1×10~5个重组子,重组率为90%,插入cDNA片段长度范围约为0．4～3kb。研究为深入开展浅色黄姑鱼的生长发育相关基因的研究奠定了良好的基础。     

黄姑鱼染色体识别与重复序列定位

黄姑鱼是我国重要的海水经济鱼类。然而,由于细胞遗传标记匮乏,黄姑鱼染色体仍然难以辨识。为了提高黄姑鱼染色体的配对识别水平,本研究利用荧光原位杂交(fluorescence in situ hybridization,FISH)、吉姆萨染色和荧光染色技术分析了黄姑鱼染色体的特征。以总DNA为探针进行基因组DNA荧光原位杂交(genomic fluorescence in situ hybridization,GISH),从而获得黄姑鱼染色体图谱,可使每对染色体呈现特定的荧光信号。依据GISH荧光信号分布模式,可以辨识黄姑鱼的24对染色体。18S r DNA FISH结果显示,18S r DNA只有一对信号,分布于1号染色体臂间,并与吉姆萨染色呈现的次缢痕、DAPI阴性带和DPI染色高亮区域同位。5S r DNA有一强一弱两对信号,信号强的一对分布于1号染色体着丝粒端,信号弱的一对分布于4号染色体的远端。端粒信号在所有染色体的端部显示,但个别染色体一端信号微弱。本研究结果丰富了黄姑鱼的细胞遗传标记,为解决黄姑鱼染色体辨识问题提供参考依据,也为进一步研究石首鱼科染色体进化提供了资料。     

日本黄姑鱼胚胎发育及温度对其过程的影响

用Nikon显微镜观察了日本黄姑鱼胚胎发育全过程,并比较了受精卵在13种温度下胚胎的发育状况。结果表明：在水温为20～22.5℃、盐度29.9和pH 8.2条件下,从受精卵至孵化出膜总历时34 h 55 min,胚胎发育过程可划分为21期;孵化水温对受精卵的孵化率、畸形率和孵化时间有显著影响,孵化时数（h）、孵化积温（A）与平均水温（θ）密切相关,h=4265.9θ-1.547 9,A=3.420 1θ2-168.99θ＋2 786.1;有效积温为480.04～553.96℃.h,阈温度为7.01℃,但实际孵化水温须高于12℃,14～28℃胚胎均能正常孵化。适宜孵化水温为17～22℃,孵化率高于89%,畸形率低于16%。     

黄姑鱼一龄幼鱼形态性状对体重的影响分析

随机选取一龄黄姑鱼幼鱼362尾,测量其体重、全长、体长、体厚、体高、头长、尾柄长、尾柄高、吻长、吻尖至背鳍起点长、眼径、眼间距共12个形态学指标。采用相关分析和通径分析方法,揭示了影响黄姑鱼一龄幼鱼体重的主要外部形态性状。结果表明,各形态性状与体重的相关系数均达到极显著水平;影响黄姑鱼一龄幼鱼体重的主要性状是体长、体高、体厚、尾柄高和眼间距;体长对体重的通径系数最大（0.400＊＊）,对体重的决定系数最高（16.00%）;眼间距对体重的直接作用较小（0.066＊＊）。主要是通过间接作用来影响体重（0.627 6）;决定分析的结果和通径分析的变化趋势一致;各形态性状对体重总的决定系数∑d=0.904,它与相关指数R2的数值相等;通过逐步回归分析方法,经偏回归系数显著性检验,建立了体质量为因变量,体长、体高、体厚、尾柄高和眼间距为自变量的多元回归回归方程为：y=-104.906＋4.336 x1＋11.654 x2＋14.015 x3＋7.138 x4＋8.761 x5。     

黄姑鱼精子的超低温冻存及细胞结构损伤的检测

以HBSS溶液为稀释液,DMSO为抗冻剂,0.25 mL麦细管为冻存管,两步降温法超低温冻存黄姑鱼精子,并用单细胞凝胶电泳(SCGE)技术检测了冻精的DNA损伤,荧光双染色流式细胞仪技术(FCM)检测了冻精的细胞膜性结构损伤。结果表明,DMSO质量分数在5%~20%时,冻精的活力与鲜精相比无显著差异;其中DMSO质量分数在10%时,冻精的激活率、运动时间及寿命分别为85.25%±3.95%、(3.23±0.27) min及(3.83±0.33) min。DMSO质量分数在25%、30%时,冻精的活力显著下降。SCGE检测显示,DMSO质量分数在5%~15%时、冻精的DNA损伤与鲜精相比差异不显著,DMSO质量分数为20%、25%、30%时,冻精的DNA损伤明显加重,冻精的DNA损伤与抗冻剂DMSO的质量分数成正相关。FCM检测显示,DMSO质量分数在5%~20%时,冻精中线粒体及细胞膜结构保持完整性的精子比例与鲜精相比无显著差异,DMSO质量分数在25%、30%时,冻精中的线粒体及细胞膜结构保持完整性的精子比例明显下降。分析认为,较高质量分数的DMSO是引起冻精活力下降,DNA、线粒体及细胞膜结构损伤加重的主要原因。     

Effect of replacing fish meal with soybean meal on growth, feed utilization and carcass composition of cuneate drum (Nibea miichthioides)

The effect of replacing fish meal with soybean meal (SBM) in practical feeds for cuneate drum was evaluated in an 8-week net pen trial. The cuneate drum fingerlings (initial body weight 29.8 ± 1.3 g fish^− 1) were fed six isonitrogenous and isocaloric feeds containing 39% digestible protein and 16 MJ kg^− 1 digestible energy. The control feed was formulated to contain 40% herring meal, whereas in the other five feeds SBM was included at 11.3, 22.5, 33.8, 45.0 and 56.3% to replace 20, 40, 60, 80 or 100% of the fish meal. There were no significant differences in feed intake between fish fed the control feed and feeds in which SBM replaced 20 to 80% of the fish meal, but fish fed the fish meal free feed had higher feed intake than the other treatments. Weight gain linearly declined with the decrease of fish meal level. Final body weight (FBW) of fish fed the feeds in which SBM replaced 20% of the fish meal did not significantly differ from fish fed the control feed. Replacing 40 to 100% of the fish meal resulted in lower FBW and nitrogen retention efficiency (NRE), and higher feed conversion ratio (FCR) than those of fish fed the control feed. Fish fed the feeds in which SBM replaced 60 to 100% of the fish meal had lower condition factor and hepatosomatic index than those of fish fed the control feed. No significant differences in carcass protein content was found among the treatments, but fish fed the feeds in which SBM replaced 60 to 100% of the fish meal had higher moisture and lower lipid content in carcass than those of fish fed the control feed. Results of the present study appear to indicate that cuneate drum has a limited ability to utilize SBM as a protein source in practical feeds.