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大黄鱼、鮸鱼及美国红鱼线粒体DNA的Cyt b基因序列比较 总被引：1，自引：0，他引：1
利用PCR技术对大黄鱼Pseudosciaena crocea、鮸鱼Scioenops ocellatus和美国红鱼Miichthys miiuy线粒体DNA的细胞色素b（Cytb）基因片段进行了扩增，PCR产物直接测序，分别获得大黄鱼和美国红鱼1140bp的序列，鮸鱼1125bp的序列。通过对3种鱼的cnb序列的比对分析，得出3种鱼序列的相似性为91．49％；分析了3种鱼序列的碱基组成及碱基变异情况，发现3种鱼的序列差异明显，碱基替换较多，可以将cytb基因作为种间分子鉴定或更高分类界元遗传分析的分子标记；计算了3种鱼序列的遗传距离并构建了系统树，结果显示，鮸鱼与美国红鱼的遗传距离较近。 相似文献
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利用PCR技术克隆唐鱼（Tanichthys albonubes）β-actin基因。所克隆的β-aetin基因片段为1464bp，包含长为1374bp的启动调控区和90bp的部分开放阅读框。启动调控区包括105bp的β-actin基因上游调控序列、第一个外显子和第一个内含子。上游调控序列中含有对转录起重要作用的CAAT Box、TATA Box、CArG Box等元件。将唐鱼β-actin启动调控区克隆到红色荧光表达载体pDsRed2-1上，并显微注射到唐鱼受精卵中，荧光显微镜观察红色荧光蛋白（RFP）的表达。结果表明，RFP在转基因唐鱼中的表达阳性率较高，最高可达51．8％，且RFP的表达水平较高。PCR检测转基因唐鱼的部分器官组织，在被检组织器官中均能检测到外源RFP基因；而RT-PCR以及Southern blot验证显示RFP mRNA的表达有所不同；Southern blot检测肌肉组织基因组DNA，可见比阳性载体大的杂交条带。说明所检测的组织中已发生外源基因RFP的整合，但有些组织存在表达水平较低或者不表达的现象。本实验分离到的β-actin基因启动子序列具有有效的驱动功能，可启动外源基因在唐鱼体内的高效表达，从而为下一步进行功能基因的转化研究奠定基础。 相似文献
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从皱纹盘鲍雌性个体的足部肌肉提取总DNA后，通过聚合酶连式反应（PCR）技术扩增得到一个扩增产物。经克隆、筛选、确定重组子产物。测序得到了长度为511bp的启动子片段。分析测序结果发现，皱纹盘鲍肌动蛋白基因启动子DNA序列与目前己知的红鲍相应序列的相似度为95%；GC碱基含量为38.93%，较红鲍的低（59.2%）；所得序列含有高度保守的基本表达调控元件，即一个CAAT框和四个TATA框。 相似文献
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大黄鱼mtDNA ND5和Cytb基因的克隆与序列分析 总被引：5，自引：0，他引：5
2004年4月，将采自浙江省象山港海区网箱养殖的大黄鱼样本，提取总DNA，通过设计特异性引物对大黄鱼线粒体DNA（mtDNA）的辅酶5（ND5）和细胞色素b（Cytb）基因进行PCR扩增。扩增产物经琼脂糖电泳检测、纯化后直接测序。得到ND5的序列1839 bp和Cytb基因序列382 bp。应用primer premier5和MEGA3软件包所作的系统发育分析表明：依据大黄鱼ND5序列所作的进化树总体支持传统的分类地位，大黄鱼更接近塘鳢鱼科。而基于Cytb基因所作的分析表明，黑鳃梅童鱼是大黄鱼在石首鱼科中是遗传距离最小的。 相似文献
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为了评估转红色荧光蛋白(red fluorescent protein,RFP)基因唐鱼新品系的稳定性,研究了RFP基因在不同世代转基因唐鱼中的遗传和表达情况。荧光显微镜下观察显示,RFP基因在F6和F10代所检测的组织器官中均有表达,并且两个世代间相同组织部位的表达水平相似。F6和F10代个体分别配对繁殖实验表明,RFP基因在转基因唐鱼后代中的遗传仍然符合孟德尔分离规律,而且培育出的转基因个体表型特征无显著差异。利用PCR技术在F2、F6和F10代转基因唐鱼基因组中扩增外源性肌球蛋白轻链2启动子、RFP基因编码区和整合位点上下游侧翼区域(片段总长度为4 883 bp),测序结果显示,3个世代间的外源基因序列完全相同,没有发生碱基缺失或突变等现象。研究表明,红色荧光蛋白基因在转基因唐鱼传代培育过程中保持了稳定的遗传和表达。 相似文献
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养殖大黄鱼病原弧菌多重PCR检测技术的建立和应用 总被引：3，自引：0，他引：3
溶藻弧菌(Vibrio alginolyticus)、副溶血弧菌(vibrio parahaemolyticus)和哈维氏弧菌(Vibrio harveyi)是浙江省养殖大黄鱼(Pseudnosciaena crocea)弧菌病的主要致病菌.本研究选择针对溶藻弧菌和副溶血弧菌的胶原酶基因,哈维氏弧菌的部分ToxR基因的特异性,优化设计了3对特异性引物,通过进行多重PCR反应体系优化,多重PCR产物的测序鉴定与特异性和敏感性实验,建立了一种检测致病性弧菌的多重PCR检测方法.经过琼脂糖凝胶电泳后的条带分析判断,可以在一个PCR管中同时成功地检测这3种病原细菌,含溶藻弧菌、哈维氏弧菌和副溶血弧菌3种致病弧菌核酸的阳性对照样品分别扩增出大小为737 hp、382 bp和271 bp的预期产物,其灵敏度是102～102 CFU/mL.将该方法应用于检测人工感染后的养殖大黄鱼病鱼肝脏和肾脏,结果在6份组织样本中,5份检出原始感染菌株,与API 20E鉴定结果相符;对弧菌病流行季节采集的未发病的16份养殖大黄鱼组织样本和16份水体样本进行抽检,结果在1份大黄鱼组织样本中检出哈维氏弧菌,7份水体样本中检出这3种弧菌中的1种或2种,鉴定结果与API 20E鉴定结果符合率为93.75%.说明该方法不仅可以检测发病鱼,还可以检测无病症带菌大黄鱼以及带菌水样,且说明海洋水体中存在着大黄鱼弧菌病的致病菌.结果说明,多重PCR检测方法具有较高的敏感性与特异性,可以缩短检测时间,降低检测成本,该方法的建立对养殖大黄鱼弧菌病的快速诊断和分子流行病学的调查具有重要意义. 相似文献
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近年来,随着肉制品掺假问题日益突出,特别是鱼肉制品中掺入廉价成分,混淆鱼肉品种,以次充好的掺假造假问题引起广泛关注。为解决这一问题,有必要针对常见的青鱼(Mylopharyngodon piceus)、草鱼(Ctenopharyngodon idellus)、鲢(Hypophthalmichthys molitrix)和鳙(Aristichthys nobilis)四大家鱼建立快速同时检测鱼肉制品的多重PCR方法。实验通过对青鱼D-LOOP基因、草鱼COI基因、鲢16SrRNA基因和鳙12SrRNA基因序列进行对比分析,针对特异序列设计4对引物,对特异基因进行单一PCR和多重PCR扩增,根据其特异性、敏感性建立并摸索L9(34)正交试验单管多重PCR最佳反应条件,包括引物浓度、Tm值、Mg2+浓度和模板量等,通过对PCR条件的优化,建立快速检测青鱼、草鱼、鲢和鳙四大家鱼鱼肉制品的单管多重PCR鉴定方法。结果表明,针对青鱼、草鱼、鲢和鳙4种鱼肉制品所设计的4对特异引物分别能扩增出298、210、306及639 bp的目的条带,具有高度特异性,反应条件优化后,4种鱼肉制品所含靶基因质粒在50 pg均可同时扩增出较清晰条带,以人工自制鱼肉制品进行了验证,结果表明所建立的方法,能够区分混合样品中4种鱼肉的构成,并对市售鱼肉制品进行检测,结果稳定。本研究所建立的方法能够简单、快速地对青鱼、草鱼、鲢和鳙4种鱼肉制品成分进行检测和监控,且特异性强、灵敏度高,可为鱼肉制品质量安全提供技术支持。 相似文献
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为探索鱼类转铁蛋白基因tf和转铁蛋白受体基因tfr1a的转录调控机制,本实验以团头鲂为研究对象,在其全基因组数据库中获取tf和tfr1a基因序列,对2个基因候选启动子区转录因子结合位点及CpG岛进行预测,通过PCR方法克隆得到tf和tfr1a基因近端启动子区不同长度片段,连接至pGL3-Basic/pEGFP-1载体,瞬时转染入Hela细胞,并采用双荧光素酶报告基因检测系统进行检测。结果发现,团头鲂tf基因启动子区无CpG岛位点,而tfr1a基因启动子区有2个CpG岛位点。成功构建9个tf和10个tfr1a不同长度启动子片段的重组质粒,经双荧光素酶报告基因系统检测发现,tf启动子核心区域为-268^+56 bp,且-1 308^-1 102 bp片段可能存在正调控该基因表达的转录因子结合位点;tfr1a启动子核心区域为-224^+48 bp,且+48^+92 bp可能存在抑制该基因转录的负调控元件,而-1 229^-1 219 bp区域可能存在促进tfr1a基因表达的正调控转录因子结合位点。 相似文献
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拟油壶菌病(Oplidiopsis disease)是海上栽培紫菜(Porphyra sensu lato)的主要病害之一,常引起紫菜大面积病烂并造成严重经济损失。本研究利用拟油壶菌感染海区内不同健康状态下的条斑紫菜(Neopyropia yezoensis)[未发生任何病烂(PyOlpH)、部分紫菜发生病烂(PyOlpM)和发生严重病烂(PyOlpS)],分析其附生菌群多样性、群落结构和主要类群之间的相互作用。结果显示,3种紫菜附生菌群α多样性指数不存在显著差异,但PyOlpM组指数高于其他组。3种紫菜附生菌群共有可操作分类单元(operational taxonomic unit, OTU)数仅占总OTU数的22.7%,菌群之间存在显著差异(置换多元方差分析, R2=0.405, P<0.05)。紫菜感染程度越高,与PyOlpH之间的差异类群数量越多。共注释出23门208属,α-变形菌纲(α-Proteobacteria)、γ-变形菌纲(γ-Proteobacteria)和厚壁菌门(Firmicutes)在所有样品中均占优势,相对丰度前20个属中有16个也位于这3个类群中,且随着感染程度的增加分别出现递增或递减的趋势。其中,贪铜菌属(Cupriavidus)和鞘氨醇单胞菌属(Sphingomonas)是共现网络中连接度最高的细菌类群,二者及其依靠正相互作用连接的细菌类群之间存在负相互作用。本研究可为阐明拟油壶菌致病的微生态机制及寻找生防细菌提供一定的数据支持。 相似文献
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近年来,我国北方沿海海参池塘养殖发展迅速,给多年来一直不景气的池塘养殖带来了新的生机,胶州湾沿岸有条件的池塘在政府的引导和政策的扶持下,纷纷改造用于养殖海参,但由于池塘老化、清淤困难,改造成本高,海参的成活率低、生长速度慢、病害多,养殖效果不甚理想,在一定程度上也 相似文献
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Caroline Stanley Maura Hiney Deirdre Gilroy Richard Powell David Padley Pete Smith 《Aquaculture (Amsterdam, Netherlands)》2002,210(1-4):35-48
Two laboratory microcosms, constructed of sterilised sediment and water taken from different locations, were seeded with Aeromonas salmonicida. A non-quantitative PCR, a heterogeneous, non-competitive, sandwich ELISA and colony formation were employed to generate survival data. In one microcosm, all three analytical methods generated positive results over the full 279 days of the incubation of the system at 18 °C. In the second microcosm, constructed with humic acid-rich material, colony formation was undetectable after 1 day but the PCR and ELISA methods generated positive results for up to 269 days.
Direct injection into fish of material from the two microcosms both seeded with a passaged strain of A. salmonicida was used to determine the disease-producing potential of the bacteria in these systems. A direct correlation was observed between the presence of colony-forming units and the ability to produce furunculosis in test fish. None of the samples that generated positive PCR and ELISA signals but in which colony formation was not detected proved capable of inducing disease in fish. The significance of these data for the application of proxy methods in epizootiological studies is discussed. 相似文献
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两种模式高密度养殖乌鳢的效益比较 总被引：1，自引：0，他引：1
一、常规养成模式及效益分析连云港市地处黄海之滨的海州湾，海产小杂鱼资源丰富，淡水养殖面积４２万亩，河沟纵横，水库星罗密布，其中精养池塘面积１万亩，为乌鳢养殖的发展提供了优越的条件。从１９９７年的几亩至２００１年３２００亩，几年的发展，形成了利用本地区丰富的海产小杂鱼资源、单放大规格鱼种进行高密度养成商品鱼的模式，这种模式将经济价值低的海水小杂鱼在短周期内转变为高档的乌鳢，解决了乌鳢作为主养鱼类的饵料问题，取得了显著的经济效益和社会效益。１．养成管理及收获一般池塘面积１亩（最大的７亩），清整后，于… 相似文献
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In order to find an efficient method for marking fish a tagging experiment has been carried out at the Fish Breeding Experimental Station, Sunndalsøra, Norway. Three different methods have been tried: jaw-tagging, cold-branding and tattooing with two different types of dye. During the winter 1972/1973 relatively large numbers of rainbow trout (12 602), sea trout (2 065) and salmon (13 141) were marked using the three marking methods mentioned. It was found that cold-branding with liquid nitrogen was the most rapid and inexpensive method for marking salmonids; after about one year between 87 and 92% of the fish can be identified.For cold-branding rainbow trout and sea trout an exposure time of 3 sec is too long.Tattooing with fluorescent pigments gave poor results in this experiment but could be a useful method for marking salmonids if a device could be constructed that tattoos bigger spots.Fast blue is not recommended for marking fast-growing salmonids. 相似文献
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Jatobá A Vieira Fdo N Buglione-Neto CC Mouriño' JL Silva BC Seiftter WQ Andreatta ER 《Fish physiology and biochemistry》2011,37(4):725-732
The aim of this study was to assess the effect of a probiotic (Lactobacillus
plantarum) supplemented diet on Nile tilapia (Oreochromis
niloticus) in a polyculture system with marine shrimp (Litopenaeus vannamei) as regards culture performance, hematology, and gut bacterial microbiota. Ten 20-m2 pens were arranged in one earthen pond
and stocked with 2 fish (41.9 g) m−2 and 10 shrimp (2.3 g) m−2, in total of 40 Nile tilapias and 200 shrimp per experimental unit. Tilapia groups in five of the experimental units were
fed a commercial diet supplemented with L. plantarum and the other five with an unsupplemented commercial diet (control). After 12 weeks of culture, the tilapia groups fed the
probiotic-supplemented diet presented values 13.6, 7.5, and 7.1% higher for feed efficiency, yield, and final weight, respectively.
Viable culturable heterotrophic bacteria counts were reduced, and the number of lactic acid bacteria was increased in the
gut of fish and shrimp fed the probiotic-supplemented diet. Hematological analyses showed higher number of thrombocytes and
leukocytes in tilapia fed the supplemented diet. L. plantarum utilized in this study colonized the gut of tilapia and shrimp and resulted in reduced number of total bacteria and increased
tilapia final weight and feed efficiency. 相似文献
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Michael R. Donaldson Robert Arlinghaus Kyle C. Hanson & Steven J. Cooke 《Fish and Fisheries》2008,9(1):79-105
Catch‐and‐release (C&R) angling is widely practised by anglers and is a common fisheries management strategy or is a by‐product of harvest regulations. Accordingly, there is a growing body of research that examines not only the mortality associated with C&R, but also the sublethal physiological and behavioural consequences. Biotelemetry offers a powerful means of remotely monitoring the behaviour, physiology and mortality of fish caught and released in their natural environment, but we contend that its usefulness is still underappreciated by scholars and managers. In this study, we review the applications of biotelemetry in C&R science, identify novel research directions, opportunities and challenges. There are now about 250 C&R studies but only one quarter of these utilize biotelemetry. In fact, almost all of the C&R studies that have used biotelemetry have been conducted within the last decade. We found that the majority of C&R telemetry studies used either radio or acoustic telemetry, while comparatively few studies have used satellite technologies. Most C&R biotelemetry studies have been used to assess mortality rates, behavioural impairments or to evaluate the effects of displacement on fish. A small fraction of studies (<8%) have used physiological sensors despite the fact that these tools are highly applicable to understanding the multiple sublethal consequences of C&R and are useful for providing mechanistic insights into endpoints such as death. We conclude that C&R science has the potential to benefit greatly from biotelemetry technology, particularly with respect to providing more robust short‐term and delayed mortality estimates and adopting a more integrative and comparative approach to understanding the lethal and sublethal impacts of C&R. However, there are still a number of challenges including (i) the need for appropriate controls and methodological approaches, (ii) the need for accounting for tagging and handling stress and mortality, and (iii) the need for certainty in assessing mortality. However, the benefits associated with C&R biotelemetry outweigh its disadvantages and limitations and thereby offer C&R researchers a suite of new tools to enhance fisheries management and conservation. 相似文献