5个虹鳟（Oncorhynchus mykiss）群体的生化遗传分析

采用不连续聚丙烯酰胺凝胶垂直板电泳技术,测定和比较了道氏虹鳟、美国加州虹鳟、挪威虹鳟、丹麦虹鳟、芬兰虹鳟5个群体的15种同工酶约40个基因座位的遗传变异。结果表明,5个虹鳟群体的多态座位比例为43.24%~47.37%;种群平均预期杂合度为0.1917~0.2178,平均实际杂合度为0.2703~0.3291;Hardy-Weinberg 遗传偏离指数(d)为0.4100~0.5331,位点有效等位基因数(Ne)为1.4865~1.5385。χ2检验表明,5个虹鳟群体多态座位基因频率均符合Hardy-Weinberg平衡。与其它鱼类相比,5个虹鳟群体的遗传多样性均处于较高水平,表明本研究的5个虹鳟群体种质资源状况尚好。聚类分析表明,挪威虹鳟、丹麦虹鳟、美国加州虹鳟、道氏虹鳟为同一聚类支,芬兰虹鳟为另一聚类支。本研究对5个群体虹鳟养殖品种的同工酶分析,为其种质资源保护和开发以及遗传育种等方面提供了基础性资料。 关键词：虹鳟;群体;同工酶;遗传结构;遗传变异     

国内虹鳟代表性养殖群体的高通量SNP芯片检测及遗传分析

本研究旨在对国内虹鳟(Oncorhynchus mykiss)代表性养殖群体开展全基因组水平的遗传评估。利用57K单核苷酸多态性(single nucleotide polymorphism,SNP)芯片,检测了来自不同地域的6个虹鳟养殖群体样本共计48尾,包括黑龙江虹鳟、黑龙江金鳟、四川虹鳟、四川金鳟、北京虹鳟和北京金鳟,共获得有效SNP位点50201个,在中国虹鳟中的多态比例达到97.7%,表明该芯片虽然基于美国和挪威虹鳟群体设计,但对中国群体同样具有良好的适用性。各群体最小等位基因频率均值为0.240~0.267,与国外主流养殖群体相近,黑龙江虹鳟、四川虹鳟和北京虹鳟群体内遗传多样性丰富,多态位点比例为83.6%~84.9%,与国外主流养殖群体相近,而黑龙江金鳟、四川金鳟和北京金鳟,多态位点比例相对较低,在60.2%~76.9%范围内。应用6个中国虹鳟群体和2个美国虹鳟群体数据开展系统发育分析、主成分分析和群体遗传结构STRUCTURE分析,结果显示8个群体可分为3个祖源类群,其中3个金鳟群体为遗传联系较紧密的一个类群,黑龙江虹鳟和北京虹鳟为一个类群,而四川虹鳟与2个美国虹鳟群体为一个类群,部分中国养殖群体中有显著离群个体存在,表明群体遗传背景不均一。本研究表明,高密度SNP芯片在我国虹鳟养殖群体遗传分析中具有广泛的应用前景,能够为种质资源评估、本土化良种培育、制种和引种工作提供基因组水平的参考信息。     

毛蚶三个地理群体生化遗传特征分析

利用同工酶技术结合聚丙烯酰胺电泳对江苏海州湾、浙江象山港、辽宁辽东湾三个海区毛蚶群体的同工酶遗传多样性进行了研究。在三个群体的9种同工酶中,共检测到了18个基因位点,海州湾、象山港和辽东湾三个群体的多态位点比率(P.95)分别为61.11%、55.56%、64.71%,平均杂合度观测值(Ho)变化范围在0.1689~0.1770间,位点有效等位基因数分别为1.4417、1.3917、1.4219,遗传多样性较丰富。各群体普遍存在着杂合子缺失现象,近交系数(Fis)分别为0.2140、0.1232、0.1797。比较了三个群体间的遗传分化指数(Fst)和群体每代迁移数(Nm),结果表明,海州湾群体和象山港群体间遗传分化较小,Fst=0.0448;而辽东湾群体与其它两个群体间的遗传分化较大,分别为Fst=0.0755,Fst=0.0812。聚类分析结果同样表明,海州湾群体首先与象山港群体相聚,后与辽东湾群体聚类。讨论了毛蚶群体分化机制,近交系数与繁殖生物学的相关性。     

凫溪香鱼群体同工酶的生化遗传分析

香鱼(Pleccoglossusaltivelis)隶属鲑形目,香鱼科,香鱼属,为我国小型名贵经济鱼类,广泛分布在北起鸭绿江南至广西北仑河等涉海溪流江河中,也可生活在大型陆封水库中[1],为一年生洄游鱼类。近年来,由于各地捕捞强度增大、水利设施建设以及环境污染日益严重等原因,各地香鱼资     

小刀蛏群体内同工酶的生化遗传分析

采用聚丙烯酰胺凝胶不连续垂直平板电泳技术对采自舟山嵊泗海区的小刀蛏群体不同个体的苹果酸脱氢酶、苹果酸酶、细胞色素氧化酶、酯酶、异柠檬酸脱氢酶、过氧化物酶、超氧化物歧化酶和葡萄糖-6-磷酸脱氢酶8种同工酶进行了生化遗传分析,共记录了21个基因座位,37个等位基因,其中有11个基因座位(Mdh-1、Mdh-2、Me-1、Me-2、Pod-1、Pod-2、Pod-3、Cod-3、Icd-1、Icd-2和Icd-3)为多态,多态座位比例为52.38%.小刀蛏群体内具一定的遗传变异能力.     

3种虹鳟养殖群体的遗传结构及遗传多样性分析

从40个随机引物中筛选出20个,采用随机扩增多态DNA(RAPD)技术,对甘肃金鳟、道氏虹鳟和美国金鳟3个群体进行了遗传结构和遗传多样性分析。结果表明,甘肃金鳟、道氏虹鳟、美国金鳟群体多态位点百分比分别为82.79%、73.33%、77.87%;平均等位基因数分别为1.8239、1.7429、1.7887,有效等位基因数分别为1.4823、1.4281、1.4527;基因多样度分别为0.2820、0.2528、0.2675,Shannon遗传多样性指数分别为0.4219、0.3801、0.4022;以上参数表明,3个群体的遗传多样性均处于偏高的水平,且以甘肃金鳟的遗传多样性最为丰富。甘肃金鳟与美国金鳟之间的遗传距离最大,道氏虹鳟与美国金鳟之间的遗传距离最小。遗传分化指数表明,种内群体间的遗传分化程度很低,而群体内个体间的分化程度很高,变异主要来源于群体内个体间。综合分析认为,甘肃金鳟遗传多样性偏高,有进一步选育的遗传潜力;同时,群体内遗传分化程度较大,说明品种的纯度可能较低,经济性状不够稳定,需进一步选育。     

３种虹鳟养殖群体的遗传结构及遗传多样性分析

从40个随机引物中筛选出20个,采用随机扩增多态DNA(RAPD)技术,对甘肃金鳟、道氏虹鳟和美国金鳟3个群体进行了遗传结构和遗传多样性分析.结果表明,甘肃金鳟、道氏虹鳟、美国金鳟群体多态位点百分比分别为82.79%、73.33 %、77.87%;平均等位基因数分别为1.8239、1.7429、1.7887,有效等位基因数分别为1 4823、1 4281、1.4527;基因多样度分别为0.2820、0.2528 、0.2675,Shannon遗传多样性指数分别为0.4219、0 3801、0 4022;以上参数表明,3个群体的遗传多样性均处于偏高的水平,且以甘肃金鳟的遗传多样性最为丰富.甘肃金鳟与美国金鳟之间的遗传距离最大,道氏虹鳟与美国金鳟之间的遗传距离最小.遗传分化指数表明,种内群体间的遗传分化程度很低,而群体内个体间的分化程度很高,变异主要来源于群体内个体间.综合分析认为,甘肃金鳟遗传多样性偏高,有进一步选育的遗传潜力;同时,群体内遗传分化程度较大,说明品种的纯度可能较低,经济性状不够稳定,需进一步选育.     

文蛤北方种群生化遗传结构与变异的研究

采用聚丙烯酰胺凝胶电泳技术,对文蛤5个北方种群进行了生化遗传分析。结果表明:文蛤5个天然种群的多态座位比例为0.475±0.064;平均杂合度为0.212±0.026,总体上遗传多样性较好,遗传变异水平较高。在各多态座位中有78.95%的座位符合Hardy-W e inberg平衡,有65.79%的座位d值为正,34.21%的座位为近交座位,存在近交,但近交程度不高。通过聚类分析可以初步判定,这5个群体同属于一个种,种群间由于地理隔离已形成了不同的地理种群。     

史氏鲟、西伯利亚鲟及其杂交种群体生化遗传学分析

采用聚丙烯酰胺凝胶电泳法对史氏鲟、西伯利亚鲟及其正、反杂交种[即史氏鲟(♀)×西伯利亚鲟(♂)杂交种、史氏鲟(♂)×西伯利亚鲟(♀)杂交种]4个群体进行了9种同工酶表达分析。结果表明,史氏鲟的多态位点比例最低(20.00%),西伯利亚鲟及史氏鲟(♂)×西伯利亚鲟(♀)杂交种的多态位点比例最高,且相等(35.00%);群体平均杂合度期望值为0.1215～0.1744;聚类分析表明,史氏鲟、史氏鲟(♀)×西伯利亚鲟(♂)杂交种聚为同一分支,西伯利亚鲟、史氏鲟(♂)×西伯利亚鲟(♀)杂交种聚为另一分支。     

红鳍东方鲀养殖群体生化遗传变异初步研究

采用水平凝胶电泳技术对红鳍东方鲀养殖群体进行了同工酶分析。本实验采用肌肉和肝脏两种组织,分析了8种酶,检测到9个位点,在0.99水平上,PGM*和IDHP*表现为多态,MPI*、LDH*、AAT*、SOD*、MDH-1*、MDH-2*和G3PDH*均为单态,多态位点比例为0.2222,平均预期杂合度为0.0998,实际观测杂合度为0.0467,平均有效等位基因数目为1.1840,PGM*位点的遗传偏离指数为-0.0707,杂合子缺失,IDHP*位点无杂合子,遗传偏离指数为-1。     

Genetic variation measured by microsatellites among three strains of domesticated rainbow trout (Oncorhynchus mykiss, Walbaum)

Genetic variation fuels selective change in natural and captive populations. In establishing a broodstock for selective improvement, the level of genetic diversity is an important consideration because it provides an indication of the scope for selective progress. Three domesticated strains of rainbow trout, Oncorhynchus mykiss (Walbaum), were examined at nine polymorphic microsatellite loci to assess detectable levels of allelic diversity and heterozygosity within and differentiation among the strains. A total of 126 alleles were observed to segregate into unique multilocus genotypes for each of the 152 individuals assayed. There was an average heterozygosity of 71.5% at these nine loci, and an average of 14 alleles at a locus. Each locus was represented by alleles unique to at least two of the three strains. Deviations from Hardy–Weinberg expectations of genotype frequencies were detected in each strain. Subsequent analysis indicated sub‐structuring within strains leading to Wahlund effects that caused these deviations. Significant differences in genotype frequencies and pairwise F_ST values demonstrated that all strains were unique. The overall F_ST of 0.089 provides additional evidence of unique genetic diversity present in each strain, and agrees well with the degree of genetic variation found in rainbow trout across broad geographical ranges. The genetic diversity contributed by each population suggests that there is greater scope for selective improvement of numerous traits within a synthetic strain combining these three strains than within any individual strain.     

Effect of GnRHa injection on milt volume in recently stripped rainbow trout Oncorhynchus mykiss

In this study, the effect of gonadotropin‐releasing hormone agonist (GnRHa) injection on milt production in spent rainbow trout was investigated. On day 0, 25 newly matured male rainbow trout (Oncorhynchus mykiss) were stripped manually, and sperm quantity (vol: mL fish^?1) and quality, spermatocrit (%), sperm count (cell mL^?1), motile sperm percentage and motility duration (s) were evaluated. After stripping, fish were randomly divided into five groups: intact; sham (injected with propylene glycol as a hormone vehicle); and groups receiving 4, 8 or 16 μg kg^?1 BW of [d ‐Ala⁶ Des‐Gly¹⁰] mGnRHa. On day 7, the fish were stripped again and the same sperm characteristics as on day 0 were measured. At the beginning of the experiment, there were no significant differences in any of the sperm quantity characteristics between groups. On day 7, expressible milt volume was significantly reduced compared with day 0 (P<0.05, t‐test) in the intact and sham groups but milt quality remained the same (P>0.05, t‐test). The present study shows that GnRHa injection with a concentration as low as 4 μg kg^?1 BW after first stripping could prevent a significant reduction in milt quantity collected 7 days later without any adverse effects on sperm quality.     

Dietary yeast culture supplementation improves initial rearing of McConaughy strain rainbow trout

The addition of a yeast culture food supplement (DVAqua from Diamond V. Mills, Cedar Rapids, IA, USA) was evaluated during feeding of McConaughy strain rainbow trout Oncorhynchus mykiss. Two trials were run consecutively, starting with initial feeding, and were replicated over 2 years. The yeast culture was added to commercial trout starter at levels of 0.125 and 0.25 g kg^?1. The number of rainbow trout that died during the first 4 weeks of rearing was significantly reduced in the tanks receiving either of the yeast culture‐supplemented diets compared with the tanks receiving a control starter diet. Ending tank weights and gain were significantly and consistently greater in the tanks fed the 0.25 g kg^?1 yeast culture diet. Individual fish weights were also significantly heavier in the fish‐fed yeast culture containing diets in each year of the study at the end of 27 days. In the trials from 4 to 8 weeks postinitial feeding, mortality rates were significantly less only in the tanks receiving 0.25 g kg^?1 yeast culture supplementation. Individual fish lengths and weights at the end of the second trial were significantly greater in the tanks receiving either level of yeast culture compared with the control diet only in 1 year. Total tank weights and feed conversion were not significantly different between any of the treatments. The inclusion of DVAqua yeast culture provided significant increases in trout survival and growth through the first 4 weeks of feeding, with continued benefits for the next 4 weeks.     

Immunohistochemical localization of rainbow trout androgen receptors in the testis

SUMMARY: We examined the distribution of two rainbow trout androgen receptors (rtAR: rtAR-α and rtAR-β) in the testis immunohistochemically using a specific antibody to clarify the target cells of androgen in spermatogenesis. Positive rtAR immunoreactivity in paraffin-embedded sections was revealed using microwave treatment, and was detected in the nuclei of Sertoli cells, Leydig cells, and other interstitial cells. The presence of rtAR in Leydig cells suggested that fish androgens regulate Leydig cell activity in an autocrine fashion similar to mammalian androgens. In addition, we found that not all Leydig cells exhibited rtAR immunoreactivity in the mature testis by double staining using anti-3β-hydroxysteroid dehydrogenase (3β-HSD) antibody. Furthermore, rtAR immunoreactivity was also detected in the nuclei of spermatogonia, spermatocytes, and spermatids. The intensity of rtAR immunoreactivity in the nuclei of spermatogonia seemed to be weaker than those of spermatocytes and spermatids. These results suggested that androgens act directly on both germ cells and somatic cells in the regulation of spermatogenesis in the rainbow trout.     

The effectiveness of adenoviral vectors to deliver and express genes in rainbow trout,Oncorhynchus mykiss (Walbaum)

The efficacy of adenoviral vectors for gene delivery into fish cells, both in vitro and in vivo, was evaluated. Vectors utilized were of human adenovirus serotype 5 (Ad), which are commonly used in human clinical trials, but have not been assessed for gene delivery to fish. Because nothing is known about Ad receptors in fish, both an Ad (Ad5Luc1) with natural tropism for the coxsackie and adenovirus receptor (CAR), as well as an infectivity enhanced Ad (Ad5LucRGD) were included within this study. Gene expression was detected in cell lines using either vector. The levels seen with Ad5LucRGD were much higher than for Ad5Luc1 in most lines except CHSE-214. Transduction of CHSE-214 cells with Ad5Luc1 could be blocked with an excess of a competitive inhibitor, suggesting that these cells possess a CAR homologue thatmediates attachment of Ad, similar to that seen in mammalian cells. In vivo gene delivery was attempted by several methods, with significant expression seen only via intramuscular injection, although infection efficiency was low. Thus it was observed that several teleost cell lines are capable of being infected and one cell line expressed a human serotype adenoviral receptor homologue that aids in Ad infection. Additionally, in vivo studies indicated that muscle tissue of rainbow trout could be infected with Ad vectors, suggesting an alternative gene delivery strategy for this animal.     

Effect of dietary inclusion of lysozyme on growth performance and plasma biochemical parameters of rainbow trout (Oncorhynchus mykiss)

This study was conducted to evaluate the effect of dietary lysozyme supplementation on growth performance and plasma biochemical parameters of rainbow trout (Oncorhynchus mykiss). Graded levels of lysozyme [0 (control), 150, 300, 450 and 600 mg kg^?1 diet] were fed to fish (initial average weight 7.76 g) for 10 weeks. Dietary lysozyme supplementation regardless of inclusion level generally improved the growth, feed utilization and nutrient retention, but significant differences were mainly observed between the 450 or 600 mg kg^?1 lysozyme group and the control group. Lysozyme supplementation generally decreased the plasma aspartate aminotransferase and alanine aminotransferase (ALT) activities, but significant difference was only observed in the ALT activity between the 450 mg kg^?1 lysozyme group and the control group. In contrast, lysozyme supplementation generally increased the plasma alkaline phosphatase and lysozyme activities, but significant difference was only observed in the lysozyme activity between the 600 mg kg^?1 lysozyme group and the control group. Lysozyme supplementation generally increased the plasma antioxidant activities of enzymes (including superoxide dismutase, glutathione peroxidase and catalase) and total antioxidant capacity, but decreased the plasma malondialdehyde level. These results suggest that dietary 450–600 mg kg^?1 lysozyme inclusion may improve the growth and non‐specific immune response of rainbow trout.     

Intestinal absorption and tissue distribution of glucose and isoleucine in rainbow trout (Oncorhynchus mykiss)

The intestinal absorption and tissue distribution of D(U¹⁴C)-glucose (¹⁴C-glc), 3-O-methyl-D(U¹⁴C)-glucose (¹⁴C-OMG) and L(U¹⁴C)-isoleucine (¹⁴C-ile) were studied in rainbow trout, Oncorhynchus mykiss, using a forced-feeding technique. The appearance of radioactivity in faeces, blood, liver, white muscle and brain was monitored over 48 h. The recoveries of radioactivity 48 h postfeeding in the fish tissues studied were 36, 57 and 48% for ¹⁴C-glc, ¹⁴C-ile and ¹⁴C-OMG, respectively. White muscle and liver contained most radioactivity on the whole tissue basis. Concentrations of ¹⁴C-glc and its derivatives in the liver and the brain exceeded those of ¹⁴C-ile and its derivatives, but the reverse was found in the white muscle. These differences may reflect differences in the metabolism of glucose and isoleucine. All tissues studied showed some differences in the accumulation of ¹⁴C originating from ¹⁴C-OMG and ¹⁴C-glc. As ¹⁴C-OMG is a nonmetabolizable glucose analogue, these differences may be the result of glucose metabolism at tissue level.     

棉粕及脱毒棉粕替代鱼粉对虹鳟生长、体成分及血清生化指标的影响

用不同比例棉粕(CM)及FeSO4处理的脱毒棉粕(CM+Fe)替代饲料中0(CM0,对照组)、10%(CM10)、20%(CM20)、30%(CM30)、50%(CM50)、70%(CM70)和50%(CM50+Fe)、70%(CM70+Fe)的鱼粉,配置成8种等氮等能的饲料,喂养(9.79±0.13)g的虹鳟(Oncorhynchus mykiss)10周,每组设3个重复,研究棉粕及脱毒棉粕对虹鳟生长性能、体成分及血清生化指标的影响。结果显示:(1)对照组虹鳟增重率(WG)与CM10～CM50以及CM50+Fe组之间差异不显著,CM70组增重率、蛋白质效率(PER)与其他各组相比显著降低,饲料系数(FCR)显著升高。(2)各试验组虹鳟全鱼和肌肉粗蛋白、粗脂肪、粗灰分与对照组差异不显著;CM0～CM70组肌肉中游离棉酚含量也逐渐增加,CM50+Fe和CM70+Fe组肌肉游离棉酚含量显著低于CM50和CM70组。(3)CM70和CM70+Fe组谷丙转氨酶与谷草转氨酶活性与对照组相比显著升高,CM10～CM50和CM50+Fe组与对照组差异不显著,血清甘油三酯含量逐渐升高,血糖含量与总蛋白含量逐渐降低。结果表明,棉粕替代鱼粉的适宜比例为19.62%。     

The use of AquaMats® to enhance growth and improve fin condition among raceway cultured rainbow trout Oncorhynchus mykiss (Walbaum)

AquaMats® are a type of artificial seaweed designed to provide structure in ponds used for fish culture and as a substrate for the growth of aquatic plants and invertebrates which in turn are a source of nutrition to cultured species. In two separate tests AquaMats® were placed into raceways used to rear rainbow trout Oncorhynchus mykiss (Walbaum) to evaluate their effect on fish growth and fin condition. In the first test, the AquaMats® were placed perpendicular to the raceway length similar to a baffle design. One treatment consisted of AquaMats® that were cleaned on a regular basis, and the other treatment consisted of AquaMats® that were not cleaned throughout the test. By the end of the test no differences were found between treatments with respect to final fish weight, specific growth rate, or feed conversion ratio. The use of AquaMats® did not improve fin condition, in fact several fins measured were significantly better among control fish. In the second test AquaMats® were placed on the raceway bottom parallel to their length and to the water flow. AquaMats® were also hung from the side of the raceway to provide cover. At the conclusion of this test no differences were found between treatments with respect to final fish weight, specific growth rate, or feed conversion ratio. The placement of AquaMats® did have a transitory impact on fin condition. Mid‐way through the test, treatment fish generally exhibited longer fins compared with the controls. However, by the end of the test, these differences were no longer detectable. The results from both tests indicate that fish were not provided with additional nutrition to the extent it improved growth. However, the use of AquaMats® did make a significant, albeit transitory, impact on fin condition.