福建省实验动物学学科发展研究报告

1实验动物科学概述1.1实验动物科学的概念实验动物科学(lAbOrATOry AnIMAl SCIEnCES)是以实验动物资源研究、质量控制和利用实验动物进行科学实验的一门综合性学科。在《学科分类与代码标准》(gb/T13745-1992)中,与实验动物科学有关的学科分别是“实验动物学“、“医学实验动物学“、“比较病理学“等

Cloning and expression analysis of the MAPKK gene in the mud crab (Scylla paramamosain)

采用QrT-PCr、rACE等方法,获得了拟穴青蟹丝裂原活化蛋白激酶激酶(MAPkk)基因CdnA全长序列。该基因全长1 558 bP,开放阅读框长度为1 224 bP,编码407个氨基酸残基。同源分析显示,该基因编码的蛋白与昆虫的相似性高达70%,推测MAPkk基因在节肢动物具有较高的保守性。经荧光定量PCr检测,MAPkk基因在拟穴青蟹多个组织中有表达,且在脑神经节和卵巢中表达量较高。在拟穴青蟹卵巢发育过程中,MAPkk基因在卵巢发育期(Ⅲ期)表达量最高,发育期为卵母细胞快速生长期,推测MAPkk具有促进卵母细胞快速生长的作用。In this paper,the MAPKK( mitogen-activated protein kinase kinase) was isolated from the mud crab,Scylla paramamosain using RT-PCR and RACE methods.The obtained full-length cDNA of MAPKK w as 1 558 bp w ith an open reading frame of 1 224 bp encoding a putative peptide of 407 amino acids.By alignment,the amino acid sequence of S.paramamosain MAPKK show ed high homology w ith those of some other animals.It suggested MAPKK w as highly conservative.Real-time PCR show ed that the MAPKK gene w as expressed in various tissues,and highly expressed in brain ganglion and ovary.The MAPKK mRNA profiles during ovarian development indicated that the expression of MAPKK w as significantly high at developing stage.We inferred that MAPKK might play a stimulative role in the ovarian development of the mud crab.国家自然科学基金项目(41076081;31272632

Preparation,Identification and Purification of Antibodies Against Four GnRHs

用3-马来酰亚胺基苯甲酸琥珀酰亚胺酯(MbS)法将人工合成的4种不同来源的促性腺激素释放激(gnrH)(即章鱼gnrH、海鞘gnrH-Ⅰ、七鳃鳗gnrH-Ⅰ和七鳃鳗gnrH-Ⅲ)分别与匙孔戚血蓝蛋白(klH)偶联后作为抗原,免疫新西兰大白兔获得gnrH抗体.用间接ElISA方法检测抗血清效价,并用WESTErn blOT鉴定其特异性.制备抗原肽亲和纯化柱,纯化多克隆抗体.结果显示,4种多克隆抗体效价高达1∶500 000(体积比),并能和相应的gnrH多肽特异性结合,纯化后的抗体在SdS-PAgE显示为单一条带.通过上述方法,获得4种高效价、高纯度的gnrH抗体,为低等动物gnrH的研究提供了理论依据.Four immunogens was prepared by conjugation of four synthetic GnRH peptides(octopus GnRH,tunicate GnRH-Ⅰ,lamprey GnRH-Ⅰand lamprey GnRH-Ⅲ) and carrier protein key-hole limpet hemocyanin(KLH) respectively by MBS method.Antibody production against each immunogen was initiated in vivo by emulsifying them in Freund′s adjuvant and administering them to 2-month-old male New Zealand rabbits by hypodermal injection.The titer of antiserums was tested by indirect ELISA,and the results showed that the titer of four antibodies was 1∶500 000.The results of Western blot analysis showed that each antibody could specifically bind to the corresponding GnRH peptide.Finally,antiserums were purified by affinity purification,and the purified antibody showed a single band in SDS-PAGE.These datas suggest that four GnRH antibody with high effect and high purity were produced,and they provide an useful tool for studying GnRH of lower animal.国家自然科学基金资助项目(41076081

Immunohistochemical Study of GnRH Receptor in the Nervous Organs of the Male Swimming Crab,Portunus trituberculatus

采用MAXVISIOnTM免疫组织化学方法,以兔抗人gnrH受体的多克隆抗体,对雄性三疣梭子蟹(POrTunuS TrITubErCulATuS)的神经器官(视神经节、脑神经节和胸神经团)进行免疫组织化学定位.结果显示:gnrH受体的免疫阳性物质在雄性三疣梭子蟹视神经节、脑神经节和胸神经团的多个部位都有较为广泛的存在,定位在神经细胞的细胞质和神经髓质中,不同神经器官gnrH受体的免疫阳性强度有一定的差异.三疣梭子蟹神经器官存在gnrH受体的免疫阳性物质,为gnrH参与其神经调节作用提供了形态学依据.The immunohistochemical technique of MaxVisionTM method and human GnRH receptor antibody against rabbit were used in this study to find out the locations of immunoreactive substances in the nervous organs of the male swimming crab,Portunus trituberculatus.The results revealed that:the immunoreactive substances of GnRH receptor were widely detected in many parts of the optic ganglion,cerebral ganglion and thoracic ganglion mass of the male crab.The immunoreactive substances located in the cytoplasm of neurosecretory cells and the neuropils.In addition,there were differentions in the intensity of immunoreaction in these three nervous organs.The immunoreactive substances of GnRH receptor existed in the nervous organs could provide morphologic evidence for GnRH participating in the neuroendocrine regulation in the swimming crab.国家自然科学基金项目(40776084;41076081

Preliminary study of gonadotropin-releasing hormone receptor (GnRH receptor) in the optic ganglion of the female swimming crab (Portunus trituberculatus) by using immunohistochemical technique

【中文摘要】 为了研究促性腺激素释放激素(GnRH)受体在甲壳动物视神经节中的存在及分布情况,采用MaxVisionTM免疫组织化学方法,以兔抗人GnRH受体的多克隆抗体,对不同发育期雌性三疣梭子蟹的视神经节进行了免疫组织化学定位。结果显示,GnRH受体的免疫阳性物质在视神经节的多个部位,视神经层、视外髓、视内髓、视端髓及X器中都有较为广泛的存在,在视外髓、视内髓与X器处的神经分泌细胞中尤为明显。不同发育期的雌性三疣梭子蟹视神经节GnRH受体的免疫阳性分布位置相似,免疫阳性强度存在一定的差异。三疣梭子蟹视神经节存在的GnRH受体免疫阳性物质,为GnRH参与视神经节调节作用提供了形态学依据。 【英文摘要】 Gonadotropin-releasing hormone (GnRH) is a kind of very important and crucial hormone in neuroregulation and reproduction.In order to elucidate whether the GnRH receptor exists or not,and to investigate its locations in the crustacean-peculiar optic ganglion as well,the immunohistochemical technique of MaxVisionTM method was used in this study.The locations of immunoreactive substances in the optic ganglion of the female swimming crabs,Portunus trituberculatus were found out in different vitellogenesis stag...国家自然科学基金项目(40776084,41076081