雌核发育草鱼及亲本倍性研究

应用流式细胞术(now cytometry, FCM)和红细胞(核)体积测量法对雌核发育草鱼、普通草鱼、湘江野鲤染色体倍性进行比较研究,检测获得的子代雌核发育草鱼倍性情况.结果表明雌核发育草鱼与普通草鱼红细胞及细胞核大小无显著差异,且DNA含量一致,湘江野鲤红细胞和细胞核体积是雌核发育草鱼、普通草鱼的两倍,DNA含量是草鱼的两倍.证实人工雌核发育草鱼具有与普通草鱼相同的染色体倍性.     

四类不同倍性鲫鱼在胚胎发育期间同工酶基因表达的比较分析

用聚丙烯酰胺梯度凝胶电泳比较分析了单倍体、二倍体、三倍体和复合四倍体4类不同倍性鲫鱼以及单倍体和二倍体鲤鱼在胚胎发育时期4种同工酶(EST,LDH,MDH,SOD)酶谱。结果表明,单倍体鲫鱼和单倍体鲤鱼胚胎与各自的二倍体胚胎相比,同工酶酶谱看不出差异;天然三倍体银鲫胚胎的MDH和SOD同工酶酶谱与二倍体鲫相似,但EST和LDH同工酶比二倍体增多了酶带,有的酶带如EST5和EST6还可在鲤鱼胚胎中找到相应的表达产物,提供了天然雌核发育三倍体银鲫杂交起源的证据;复合四倍体由于含有鲤鱼的一个外来基因组,其胚胎的基因表达有些与杂种类似,在所分析的4种同工酶酶谱中,都可观察到来自鲤鱼基因的影响。此外,在由源于不同复合四倍体个体的卵子发育形成的胚胎间,还观察到同工酶基因表达的异质性。     

人工雌核发育草鱼染色体倍性的鉴定

运用经典的红细胞及细胞核体积大小测量方法以及流式细胞仪,检测了人工诱导雌核发育草鱼染色体倍性与DNA含量.雌核发育草鱼红细胞体积为(333.5±41.94)μm3,细胞核体积为(20.7±2.378)μm3;与所测普通草鱼红细胞体积(343.8±50.1)μm3,细胞核体积(21.2±1.98)μm3,没有显著差异.雌核发育草鱼DNA含量(2C)平均为2.23pg,普通草鱼DNA含量(2C)2.20pg,两者无显著差异.研究结果表明,人工雌核发育草鱼与普通草鱼具有相同的染色体倍性.     

异源精子遗传物质对雌核发育草鱼基因组的影响分析

为分析外源精子对人工诱导雌核发育草鱼基因组的影响,用随机扩增多态性和微卫星技术对经两代连续人工诱导,遗传背景一致的雌核发育草鱼以及母本草鱼和父本鲤鱼的基因组DNA进行了比较分析。10个随机引物在雌核发育草鱼中共检测到104个RAPD位点,在鲤鱼中检测到103个位点。7对微卫星引物在雌核发育草鱼中共扩增出4个微卫星位点,在鲤鱼中检测到22个位点。两种方法在雌核发育草鱼和鲤鱼中所检测到的位点均没有一个相同。根据RAPD和微卫星分析数据进行的遗传相似度分析表明二代人工雌核发育草鱼群体与其一代人工诱导雌核发育草鱼母本的遗传相似度从0.9903到1.000,与父本鲤鱼的遗传相似度为0.000。这些实验结果证明在适当的紫外线处理强度下,鲤鱼精子的遗传物质能够被完全破坏,不会对雌核发育草鱼的基因组造成遗传污染。     

高背银鲫、大口胭脂鱼及胭脂鲫(F1)不同组织的同工酶的表型分析

本文对高背银鲫、大口胭脂鱼及胭脂鲫（F1）不同组织的脂酶（EST）、乳酸脱氢酶（LDH）、苹果酸脱氢酶（MDH）、超氧化物歧化酶（SOD）四种酶的同工酶的表型进行了分析,结果表明：在肝脏中胭脂鲫EST多一条谱带,而在性腺中,LDH、MDH、SOD同工酶谱带,胭脂鲫与父母本均存在一定的差异。     

罗氏沼虾个体发育早期的同工酶研究

采用聚两燃酰胺梯度凝胶电泳技术,对罗氏沼虾个体发育早期9个时期的八种同工酶系统（EST、ALP、AMY、GDH、MDH、LDH、SOD、ME)进行研究,结果表明：SOD、ME在早期发育过程中酶谱相对稳定,SOD表现为三条谱带,ME表现为两条谱带;面EST、ALP、AMY、GDH、MDH、LDH则随发育其酶谱表现出明显差异,酶谱渐趋复杂。     

草鱼同工酶发育遗传学研究——Ⅰ.不同组织器官的同工酶分析

采用垂直淀粉凝胶电泳及特异性组织化学染色技术,研究了草鱼成体脑、眼、心、肾、肌、肝等6种组织中的6种同工酶系统(LDH、MDH、GDH、ADH、LDH、EST)的分化表达谱式。结果表明,草鱼的同工酶系统具有明显的组织特异性。与绝大多数硬骨鱼类相比,草鱼的LDH、m-MDH和ADH同工酶具有特殊的表达谱式:m-MDH和ADH均由两个基因座位编码;肾脏在LDH-A_3B与LDH-A_2B_3之间多出1条LDH酶带(LDH-X)。本文还讨论了草鱼同工酶的遗传基础和亚基组成,以及本实验的某些结果与其他作者的结果不相符的原因。     

两个雌核发育白鲢群体同工酶分析及遗传标记的确定

取源于武汉两个不同渔场两尾白鲢的卵子,经紫外照射遗传物质失活的鲤鱼精子刺激雌核发育和热休克诱导第二极体保留的基因组操作技术,获得了两个不同的人工雌核发育白鲢群体。采用聚丙烯酰胺垂直板电泳技术,分析了这两个不同人工雌核发育白鲢群体(分别称为Hy-G1和Hy-G2)内不同个体的肝脏、肌肉组织以及红细胞中乳酸脱氢酶(LDH)、苹果酸脱氢酶(MDH)、酯酶(EST)、超氧化物歧化酶(SOD)等几种同工酶的表达谱式,并与普通繁殖的同龄白鲢进行了比较。结果表明,各个雌核发育白鲢群体内不同个体间的酶谱表现出很大程度的一致性,具较高的纯合度,而两个不同雌核发育群体Hy-G1和Hy-G2之间表现有明显差异。特别是肝脏和肌肉组织迁移率较快的酯酶谱带等以其稳定的差异,建议作为区分这两个人工雌核发育白鲢群体的生化遗传标记。       

暗纹东方TUN 同工酶生化表现型的研究

利用聚丙烯酰胺凝胶垂直平板电泳方法,对暗纹东方|TUN|的心、肝、肾、肌、性腺5种不同组织的7种同工酶(EST、LDH、POD、MDH、SOD、SDH、α-AMY)进行了研究, 讨论了各同工酶的基因表达谱式,观察到EST同工酶存在着多态现象;LDH同工酶有二个基因位点,但只表现3条带,A与B亚基的结合受阻; MDH同工酶存在性别差异,说明决定MDH同工酶表达的因素在不同性别中存在差异;SOD同工酶有3个基因位点。各同工酶酶谱稳定,有组织特异性,但EST、MDH、SOD、POD同工酶在各组织器官中又表现出较大的一致性,有利于物种的鉴定。α-AMY与SDH只在个别组织中有活性,可能与特定组织与器官的形态发生与机能分化有关。 Abstract:By means of polyacrylamid gel electrophoresis seven isozymes(EST.LDH,SOD,MDH,SDH, α-AMY)in heart,liver,kidney,muscle and gonad of Fugu obscurus were studied.The gene expression patterns of each isozyme were analyzed.The results indicated that polymorphism was detected in EST isozymes;LDH isozymes had two loci,but only three bands could be observed,the random association of two subunits(A and B)were restricted;Some MDH isozymes existed sexual differences in identical tissues.The suggested that the factors controlling the expression of MDH isozymes were different between sexes.All the isozyme phenotypes exhibited tissue-specificity and stability,but EST,MDH,SOD and POD isozymes showed relatively consistence in the five tissues.Their characteristic bands could be used in species determination.The activities of SDH andα-AMY isozymes could only be detected in some tissues and closely correlated to the morphological or functional differentiation of those tissues or organs.     

草鱼和鲤杂交的细胞学研究—鱼类远缘杂交核质不同步现象

以草鱼(Ctenopharyngodon idellus)为母本、鲤(Cyprinus carpio)为父本进行人工杂交,杂种胚胎发育至孵化期全部死亡;同时获得了少数雌核发育草鱼和雄核发育的鲤。分析比较了草鲤杂种胚胎染色体变化及胚胎发育情况。发现杂种胚胎染色体数目变化较大,一般在24—73之间,绝大部分细胞染色体在发育过程中不断丢失而出现非整倍体;极少数细胞在受精后雌性原核和雄性原核不结合而引起雌核发育和雄核发育;草鱼和鲤胚胎发育时序有较大差别;因此细胞分裂不能同步。可能是杂种胚胎染色体不断丢失的主要原因。     

四类不同倍性鲫鱼在胚胎发育期间同工酶基因表达的比较分析

Isozyme zymograms of esterase (EST), lactate dehydrogenase (LDH), malate dehydrogenase (MDH) and superoxide dismutase (SOD) were analysed by polyacrylamide gradient gel electrophoresis at different developmental stages of embryogenesis in 4 types of various ploidy crucian carp embryos, including haploids, diploids, natural triploids, and multiple tetraploids, and 2 types of haploid and diploid common carp embryos. Haploid embryos of crucian carp (Carassius auratus) and common carp (Cyprinus carpio) were produced by treating eggs with UV-irradiated milt from blunt snout bream (Megalobrama amblycephala). Natural triploid embryos were obtained from the eggs of gynogenetic silver crucian carp (Carassius auratus gibelio) inseminated with milt from red common carp. Multiple tetraploid embryos were also produced by gynogenesis from eggs of the newly discovered multiple tetraploid females inseminated with milt from red common carp. Gradient gel electrophoresis indicated that the band types and staining intensity of 4 isozymes expressed in haploid embryos of crucian carp and red common carp were similar to that in the correlative diploid embryos. In natural triploid silver crucian carp embryos, the zymograms of MDH and SOD isozymes were identical with that of diploid crucian carp embryos, but the EST and LDH isozymes manifested more new enzyme bands in comparison with diploid embryos. The corresponding expressed products of some bands in the triploid embryos, such as EST5 and EST6, could be observed also in red common carp embryos, which provided evidence for hybrid origin about the gynogenetic fish. The multiple tetraploids incorporated one foreign genome of red common carp, therefore, the effects of genes from the foreign genome could be observed in the multiple tetraploid embryos. Gene expression of the isozymes in the tetraploid embryos was somewhat similar to that in hybrids. Owing to interaction of triploid silver crucian carp genomes and common carp haploid genome, some isozyme bands, such as EST5 and EST6, changed in quantity, and some bands increased, such as s-SOD1, s-SOD2, s-SOD3 and s-SOD4 in the tetraploid embryos. Moreover, the heterogeneity was revealed among embryos developed from gynogenetic eggs of 3 different multiple tetraploid individuals.     

复合四倍体异育银鲫个体间遗传异质性的研究

用聚丙烯酰胺梯度凝胶电泳,分析比较了４个不同的银鲫雌核发育系,红鲤和复合四倍体异育银鲫鳍的５种不同的同工酶及蛋白表型的差异,表明复合四倍体异育银鲫表型上的差异主要是来自银鲫种内的遗传差异（不同的雌核发育系）。来源于同一个银鲫雌核发育系的复合四倍体异育银鲫,个体间的ＥＳＴ同工酶出现差异,并与父体红鲤的ＥＳＴ同工酶的多态性相关,因此,复合四倍体异育银鲫个体间的异质性也包含了来自父本的遗传影响。在所检测     

云南高背鲫鱼不同组织同工酶分析

采用聚丙烯酰胺凝胶电泳技术,分析了滇池中云南高背鲫鱼(Carassius auratus)的脑、眼睛、肝脏、心脏、肌肉5种组织中的酯酶(EST)、超氧化物歧化酶(SOD)和乳酸脱氢酶(LDH)3种同工酶的表达模式,并对各种酶的同工酶酶谱表型进行了分析。结果表明,云南高背鲫鱼的同工酶系统具有明显的组织特异性,且同工酶的种类与活性变化与其功能相适应。     

泽蛙雌核单倍体几种同工酶基因表达的研究

本研究用聚丙烯酰胺凝胶垂直平板电泳,比较泽蛙雌核生殖单倍体和泽蛙二倍体在鳃盖闭合期的乳酸脱氢酶同工酶(LDH)、苹果酸脱氢酶同工酶(MDH)、葡萄糖-6-磷酸脱氢酶同工酶(G-6-PDH)和酯酶同工酶(EST)的表型。实验表明,单倍体的上述同工酶区带数和活力与同胚期的二倍体比较,差异甚大。由此,作者认为,泽蛙单倍体问工酶基因的表达异常是由于缺少另一套染色体基因的相互作用。     

中华姬鼠与大林姬鼠的同工酶差异

中华姬鼠（Ａｐｏｄｅｍｕｓｄｒａｃｏ）和大林姬鼠（Ａｐｏｄｅｍｕｓｐｅｎｉｎｓｕｌａｅ）是形态学上十分相似的两种鼠类。为了对两种姬鼠的分类提供生物化学方面的依据,采用聚丙烯酰胺凝胶等电聚焦电泳方法比较和分析了两种姬鼠的ＬＤＨ同工酶、ＥＳＴ同工酶和ＳＯＤ同工酶的差异。结果表明,两种姬鼠的ＬＤＨ同工酶酶谱基本相似,而ＥＳＴ同工酶和ＳＯＤ同工酶酶谱则存在明显的种间差异。根据ＥＳＴ同工酶Ａ２带的有无和ＳＯＤ同工酶主带等电点的差别,能将两种姬鼠很容易区分开来。     

Isozyme expression in F1 hybrids between carp and goldfish

Interspecific genetic differences in malate dehydrogenase (MDH), lactate dehydrogenase (LDH), superoxide dismutase (SOD), and esterase (EST) isozymes in carp (Cyprinus carpio) and goldfish (Carassius auratus) were used to examine the allelic expressions in the hybrid between these species. A unique liver SOD and muscle LDH phenotype unambiguously identifies all presumed hybrid individuals. There was no evidence of F₂ or backcross phenotypes in hybrid individuals. Liver MDH and EST phenotypes in hybrids show a preferential expression of goldfish isozymes. Variation in the levels of carp liver MDH isozymes may result from the polymorphism of a regulatory mutation affecting isozyme expression, leading to gene silencing after duplication.This work was supported through NSERC (Canada) grants to James P. Bogart and John F. Leatherland.     

社鼠组织器官同工酶的研究

用聚丙烯酰胺凝胶等电聚焦电泳方法,分析了社鼠的肝、肾、心肌、骨骼肌、肺、脾和脑等多种组织器官的ＬＤＨ、ＡＤＨ、ＥＳＴ、和ＳＯＤ４种同工酶,对各组织器官的酶带数目和分布,以及酶活性进行了比较研究。结果表明,社鼠的ＬＤＨ同工酶、ＡＤＨ同工酶和ＥＳＴ同工酶具有比较明显的组织特异性,而ＳＯＤ同工酶的组织特异性较低。肺和脾除ＥＳＴ同工酶活性较高外,脑除ＬＤＨ同工酶活性较高外,其它３种同工酶的活性均较低;而肝和肾中４种同工酶的活性普遍很高。心肌和骨骼肌因氧张力不同而使ＬＤＨ同工酶酶谱存在明显差异,但其它３种同工酶酶谱却非常相似。同工酶的组织特异性与各组织器官所执行的生理功能是相一致的