Cloning and expression of estrogen related receptor gene from Scylla paramamosain and Tachypleus tridentatus

雌激素相关受体（estrogen-relatedreceptors，ERRs）属于孤儿核受体，在结构上和雌激素受体（estrogenreceptors，ERs）非常类似，能够直接与类固醇受体共激活子结合激活靶基因的表达。对哺乳动物的研究表明，ERRs参与雌激素信号通路，与生殖和发育过程有关。 本文采用RT-PCR、RACE、GenomeWalking等方法从拟穴青蟹（Scyllaparamamosain）和中国鲎（Tachypleustridentatus）中克隆得到雌激素相关受体基因，分别命名为spERR和ttERR，并采用生物信息学方法对其序列和编码的蛋白质的理化性质、分子系统进化关系等...The estrogen related receptors ERRs, belonging to orphan nuclear receptors, share the similar structure with the estrogen receptors ERs. ERRs can bind to the steroid receptor coactivator family without any ligands and drive transcription activity of the target genes. The results have suggested mammalian ERRs collaborate with the estrogen signaling, involving in reproductive and developmental proce...学位：理学硕士院系专业：海洋与环境学院海洋学系_海洋生物学学号：2242008115148

Hepatopancreas cell cultures from mud crab, Scylla paramamosain

Hepatopancreas is an important digestive and endocrine organ in crustacean. However, there are few reports on cell cultures from crabs. Here, the cell cultures of hepatopancreas from Scylla paramamosain was studied in vitro. Both the primary cell culture and subculture were grown in Leibovitz' L-15 medium, M199 medium, or a specially designed medium for S. paramamosain (MSP). The results showed that hepatopancreas cells in vitro grew in compact clusters in 2-3 d. Four types of cells could be identified. They were embryo cells, fibrillar cells, resorptive cells, and blister-like cells, respectively. Some of these cells could be subcultured for three generations. The MSP supported the best survival of these hepatopancreas cells, while M199 medium was the least effective of these three media. Fetal bovine serum and crab muscle extracts as supplements stimulated growth, but the crab hemolymph inhibited cell growth. Taken together, MSP is an appropriate medium for hepatopancreas cell cultures from S. paramamosain and can support cultures through several passages.National Natural Science Foundation of China [40776084]; National High Technology Research and Development Program of China [2006AA10A406

Cloning and Analysis of β-actin Gene from Scylla paramamosain

β-肌动蛋白广泛存在于真核生物中,在维持细胞结构、细胞运动和细胞分裂等生理活动中发挥着重要作用.运用rACE技术克隆了拟穴青蟹(SCyllA PArAMAMOSAIn)β-肌动蛋白基因,并用rT-PCr方法检测该基因在成体各组织中的表达情况.拟穴青蟹β-肌动蛋白CdnA全长1 337bP,5′端非编码区为67bP,3′端非编码区为139bP,开放阅读框1 131bP编码376个氨基酸.拟穴青蟹β-肌动蛋白与其他节肢动物β-肌动蛋白氨基酸序列的相似性高达98%~99%.系统进化树显示拟穴青蟹β-肌动蛋白基因的分子进化地位与其生物学分类地位一致.半定量rT-PCr分析结果表明,β-肌动蛋白基因在拟穴青蟹视神经节、脑神经节、胸神经节、性腺、鳃、心、胃、肌肉、肝胰腺共9个组织器官中的表达基本一致,具有良好的稳定性.β-actin plays important roles in maintaining cytoskeletal structure,cell motility and cell division in eukaryote.In this paper,β-actin gene was cloned from Scylla paramamosain using RACE.The results showed that the full-length cDNA of β-actin gene was 1 337 bp,containing a 5′-untranslated region(5′-UTR) of 67 bp and a 3′-UTR of 139 bp.The open reading frame of 1 131 bp encodes a 376-amino acid protein.The amino acid sequence of β-actin in S.paramamosain shared high similarity with other arthropods(98%-99%).The phylogenetic analysis was consistent with traditional classification.Semi-quantitative RT-PCR analysis showed that the expression level of β-actin gene in S.paramamosain was stable in tissues:optic ganglion,brain,thoracic ganglion,ovary,gill,heart,stomach,muscle and hepatopancreas.国家自然科学基金项目(41076081

Molecular Cloning and Expression Profiles of β-actin Gene in Various Tissues of the Horseshoe Crab,Tachypleus tridentatus

中国鲎(TACHyPlEuS TrIdEnTATuS)是一种古老的海洋动物.利用rT-PCr和rACE等分子生物学技术,获得中国鲎β-肌动蛋白基因(中国鲎β-ACTIn,简称为TTbA)全长CdnA序列,总共1 515bP,包括70bP 5′非编码区(unTrAnSlATEd rEgIOnS,uTr)、314bP 3′uTr和一个1 131bP的开放阅读框(OPEn rEAdIng frAME,Orf).Orf可编码376个氨基酸残基,总分子质量约为41 807.7u.同源蛋白的序列比较结果显示,TTbA推导氨基酸序列与其他物种具有很高的相似性,推测β-ACTIn基因具有很高的遗传保守性.而基于β-ACTIn蛋白序列比对而绘制的系统进化树显示中国鲎与其他节肢动物具有较高的相似性,此结果与其现行的分类地位基本一致.以rEAl-TIME rT-PCr法检测表明,在卵子发生各期的卵巢组织中TTbA表达量保持稳定(P>0.05),可作为定量检测的内参基因.The horseshoe crab(Tachypleus tridentatus)is an ancient marine organism.In this study,a full-length cDNA ofβ-actin(designated as TTBA)was cloned from T.tridentatus using RT-PCR and RACE method.Data showed that the full-length cDNA was 1 515 bp,including a 5′untranslated region(UTR)of 70 bp,a 3′UTR of 314 bp,and an open reading frame(ORF)of 1 131 bp nucleotides.The ORF encoded a polypeptide of 376 amino acids with calculated molecular mass of 41 807.7u.Amino acid sequence alignment showed the TTBA had high similarity with that in other organisms.It suggested thatβ-actin was highly conservative.Phylogenetic analysis based on comparison ofβ-actin proteins showed that T.tridentatus was more similar to arthropod than to other invertebrate species.This result was consistent with the current evolution relationship.Moreover,gene expression profiles by Real-Time RT-PCR revealed that the abundance of TTBA mRNA kept steady during oogenesis in ovary(p>0.05)indicating that TTBAis suitable as an endogenous control for gene expression study.国家自然科学基金(40406030

碳离子诱导的DNA双链断裂

ＤＮＡ双链断裂（ＤＳＢｓ）是电离辐射诱导的最重要的原发损伤，研究ＤＳＢｓ有利于揭示细胞辐射敏感性的机理。用倒转脉冲场凝胶电泳结合荧光扫描进行ＤＮＡ定量研究７５ＭｅＶ／ｕ１２Ｃ６＋对小鼠Ｂ１６黑色素瘤细胞ＤＳＢｓ的诱导，结果表明：ＤＳＢｓ产额约为０．７４ＤＳＢｓ／１００Ｍｂｐ／Ｇｙ；ＤＮＡ片段分布在两个区域。大片段区分子量约为１．４Ｍｂｐ～３．２Ｍｂｐ，分子量小于１．２Ｍｂｐ的为小片段区；并且随着剂量的增加，大片段区ＤＮＡ含量逐渐下降，而小片段区的ＤＮＡ含量显著增加。表明Ｂ１６ＤＮＡ分子上可能存在对重离子较为敏感的位点。攀登计划（B）”项目,中国科学院兰州分院基

JUNO Sensitivity on Proton Decay p→νˉK+p\to \bar\nu K^+ Searches

The Jiangmen Underground Neutrino Observatory (JUNO) is a large liquid scintillator detector designed to explore many topics in fundamental physics. In this paper, the potential on searching for proton decay in $p\to \bar\nu K^+$ mode with JUNO is investigated.The kaon and its decay particles feature a clear three-fold coincidence signature that results in a high efficiency for identification. Moreover, the excellent energy resolution of JUNO permits to suppress the sizable background caused by other delayed signals. Based on these advantages, the detection efficiency for the proton decay via $p\to \bar\nu K^+$ is 36.9% with a background level of 0.2 events after 10 years of data taking. The estimated sensitivity based on 200 kton-years exposure is $9.6 \times 10^{33}$ years, competitive with the current best limits on the proton lifetime in this channel

JUNO sensitivity on proton decay p → ν K + searches*

The Jiangmen Underground Neutrino Observatory (JUNO) is a large liquid scintillator detector designed to explore many topics in fundamental physics. In this study, the potential of searching for proton decay in the $p\to \bar{\nu} K^+$ mode with JUNO is investigated. The kaon and its decay particles feature a clear three-fold coincidence signature that results in a high efficiency for identification. Moreover, the excellent energy resolution of JUNO permits suppression of the sizable background caused by other delayed signals. Based on these advantages, the detection efficiency for the proton decay via $p\to \bar{\nu} K^+$ is 36.9% ± 4.9% with a background level of $0.2\pm 0.05({\rm syst})\pm$$0.2({\rm stat})$ events after 10 years of data collection. The estimated sensitivity based on 200 kton-years of exposure is $9.6 \times 10^{33}$ years, which is competitive with the current best limits on the proton lifetime in this channel and complements the use of different detection technologies