大黄鱼(Larimichthys crocea)补体C3和C4基因的分子特征及表达分析     

王海玲  祁鹏志  郭宝英  吴常文 《海洋与湖沼》2015,46(1):181-190

本文测定了大黄鱼C3(L.c-C3)和C4(L.c-C4)基因的c DNA全序列。结果表明,L.c-C3和L.cC4序列全长分别为4962bp和5088bp,分别编码1653和1695个氨基酸,N端信号肽序列分别为23和19个氨基酸。推导的氨基酸序列结构分析表明大黄鱼C3和C4与已报道的补体C3、C4同样都具有在功能上比较重要的残基以及保守的硫酯区。分子进化分析表明,L.c-C3和L.c-C4分别与鮸鱼C3、C4的氨基酸同源性最高。实时荧光定量PCR结果显示,L.c-C3和L.c-C4在健康大黄鱼的肝脏、脾脏、肠、鳃、心脏、脑、肌肉和胃这8种组织中都有表达,其中肝脏的表达量最高。在大黄鱼胚胎不同发育时期(从2细胞期到初生仔鱼)中,L.c-C3在各个阶段没有明显的变化,而L.c-C4的表达量有明显升高。溶藻弧菌(Vibrio alginolyticus)侵染的大黄鱼肝脏和脾脏中,L.c-C3和L.c-C4的m RNA表达量均明显上调。该结果表明,大黄鱼肝组织C3和C4基因表达变化与溶藻弧菌的侵染密切相关,揭示了C3和C4在大黄鱼抗细菌免疫反应中具有重要的作用。  相似文献   

鳗弧菌(Vibrio anguillarum)侵染对青蛤(Cyclina sinensis)髓样分化因子88基因表达的影响     

高晶  任毅鹏  潘宝平  闫春财 《海洋与湖沼》2015,46(2):440-445

经转录组测序后筛选并克隆得到青蛤(Cyclina sinensis)髓样分化因子88(myeloid differenttiation factor 88,My D88)的c DNA序列。在鳗弧菌(Vibrio anguillarum)胁迫下,采用荧光定量PCR法分析了My D88基因在青蛤体内的表达过程。结果显示,青蛤My D88基因的开放阅读框为1521bp,编码506个氨基酸,分子量约为57.14k Da,氨基酸N段存在DEATH结构域,C段存在TIR结构域(Toll/Interleukin-1 receptor domain)。My D88基因在青蛤血淋巴、肝脏、外套膜、鳃和闭壳肌等组织中普遍表达,但在血淋巴中表达量最高,与其它组织出现显著性差异(P0.05)。通过检测鳗弧菌刺激下My D88基因在青蛤血淋巴中的表达值,发现My D88基因在24h开始升高,48h达到最大值,约为对照组的10倍,实验组与对照组及空白组均出现了极显著性差异(P0.01);研究结果表明,该基因在软体动物的免疫应答反应中对革兰氏阴性菌有识别作用。  相似文献   

牙鲆(Paralichthys olivaceus)TLR21 基因在迟缓爱德华氏菌(Edwardsiella tarda)感染后的表达特征     

张洁  郑津辉  李庆亚  耿绪云  孙金生  潘宝平  孙世南  高虹 《海洋与湖沼》2015,46(6):1502-1508

应用荧光定量PCR技术, 检测了TLR21 基因在牙鲆(Paralichthys olivaceus)感染迟缓爱德华 氏菌(Edwardsiella tarda)后, 在0 h、1 h、3 h、6 h、12 h、1 d、3 d、6 d 后, 在心脏、肝脏、脾脏、 头肾、鳃、小肠、肌肉和血的时空表达特征, 并探讨了它们与牙鲆先天免疫反应的关系。结果表明, 大 多组织在感染病原6 h 后TLR21 基因表达明显上调, 尤其是头肾和小肠。头肾6 h 的表达量达到了 对照组的59.3 倍, 小肠6 h 的表达量为对照组的38.6 倍。迟缓爱德华氏菌感染引起牙鲆体内各组织 中TLR21 的上调表达和变化, 为研究牙鲆对迟缓爱德华氏菌的防御机制提供了理论依据。  相似文献   

长牡蛎(Crassostrea gigas)微卫星多重PCR 体系构建及其在家系鉴定中的应用     

纪仁平  李焕军  冯艳微  张荣良  王卫军  杨建敏 《海洋与湖沼》2015,46(6):1542-1548

筛选多态性高、特异性好、片段大小适中的10 个长牡蛎微卫星位点进行优化组合, 根据扩 增片段大小及同一荧光不重叠原则构建了两组五重PCR 体系。运用CERVUS 3.0 软件对0527 组27 个长牡蛎全同胞家系的643 个子代和0612 组27 个全同胞家系382 个子代分别进行亲权鉴定。结果 发现, 用两组微卫星五重PCR 鉴别时, 在0527 组和0612 组的鉴定成功率均为100%; 只用第一组微 卫星五重PCR, 可以将0527 组96%的子代和0612 组96%的子代鉴定到亲本; 只用第二组微卫星五 重PCR, 可以将0527 组97%的子代和0612 组95%的子代鉴定到亲本。本研究中筛选出的两组微卫 星五重PCR 体系在两组家系中的鉴定效率均较高, 可以快速有效地将子代个体鉴定至所属父母本, 在长牡蛎家系鉴定中具有很好的应用价值。  相似文献   

Depuration dynamics of oysters (Crassostrea gigas) artificially contaminated by Salmonella enterica serovar Typhimurium     

de Abreu Corrêa A  Albarnaz JD  Moresco V  Poli CR  Teixeira AL  Oliveira Simões CM  Monte Barardi CR 《Marine environmental research》2007,63(5):479-489

The State of Santa Catarina produces the greatest quantity of edible mollusks in Brazil. To guarantee sanitary qualify, mollusk cultures should be monitored for contamination by pathogenic microorganisms. A self-purification or "depuration" system that eliminates Salmonella enterica serovar Typhimurium contamination from oysters has been developed and evaluated. The depuration process occurred within a closed system, in which 1000 L of water was recirculated for 24 h. The water was sterilized with ultraviolet (UV) light, chlorine, or both together. Oysters (Crassostrea gigas) artificially contaminated with S. typhimurium were harvested every 6 h. Samples of oyster tissue were excised and both the presence and numbers of bacteria were determined. Combined UV light and chlorine treatments resulted in total elimination of bacteria within 12 h. Polymerase chain reaction detected bacteria in water exposed to the three treatments. This pioneering study is the first of its kind in Brazil and represents a major contribution to commercial mollusk culture in this country.  相似文献   

Multiplex PCR allows simultaneous detection of pathogens in ships' ballast water     

Aridgides LJ  Doblin MA  Berke T  Dobbs FC  Matson DO  Drake LA 《Marine pollution bulletin》2004,48(11-12):1096-1101

There is enormous potential for global transfer of microorganisms, including pathogens, in ships' ballast water. We contend that a major advancement in the study of ballast-water microorganisms in particular, and of aquatic pathogens in general, will be expedited sample analysis, such as provided by the elegant technology of DNA microarrays. In order to use DNA microarrays, however, one must establish the appropriate conditions to bind target sequences in samples to multiple probes on the microarrays. We conducted proof-of-concept experiments to optimize simultaneous detection of multiple microorganisms using polymerase chain reaction (PCR) and Southern hybridization. We chose three target organisms, all potentially found in ballast water: a calicivirus, the bacterium Vibrio cholerae, and the photosynthetic protist Aureococcus anophagefferens. Here, we show simultaneous detection of multiple pathogens is possible, a result supporting the promising future use of microarrays for simultaneous detection of pathogens in ballast water.  相似文献   

Analysis of the 16S-23S rDNA intergenic spacers (IGSs) of marine vibrios for species-specific signature DNA sequences   总被引：7，自引：0，他引：7  

Lee SK  Wang HZ  Law SH  Wu RS  Kong RY 《Marine pollution bulletin》2002,44(5):412-420

Vibrios are widespread in the marine environment and a few pathogenic species are known to be commonly associated with outbreaks of diarrheal diseases in humans due to the consumption of raw or improperly cooked seafood. However, there are also many Vibrio species which are potentially pathogenic to vertebrate and invertebrate aquatic animals, and of which little is known. In an attempt to develop rapid PCR detection methods for these latter class of vibrios, we have examined the 16S-23S intergenic spacers (IGSs) of 10 lesser-known Vibrio species and successfully developed species-specific primers for eight of them--Vibrio costicola, V. diazotrophicus, V. fluvialis, V. nigripulchritudo, V. proteolyticus, V. salmonicida, V. splendidus and V. tubiashii. The IGS amplicons were amplified using primers complementary to conserved regions of the 16S and 23S rRNA genes, and cloned into plasmid vectors and sequenced. Analysis of the IGS sequences showed that 37 ribosomal RNA (rrn) operons representing seven different IGS types have been cloned from the 10 vibrios. The three IGS types--IGS(0), IGS(IA) and IGS(Glu)--were the most prevalent forms detected. Multiple alignment of representative sequences of these three IGS types from different Vibrio species revealed several domains of high sequence variability, which were used to design species-specific primers for PCR. The specificity of the primers were evaluated using total DNA prepared from different Vibrio species and bacterial genera. The results showed that the PCR method can be used to reliably detect eight of the 10 Vibrio species in marine waters in this study.  相似文献   

A Method To Detect Viable Helicobacter pylori Bacteria in Groundwater     

Debbie Flanigan  Mark Rodgers 《洁净——土壤、空气、水》2003,31(1):45-48

The inability to detect the presence of viable Helicobacter pylori bacteria in environmental waters has hindered the public health community in assessing the role water may play in the transmission of this pathogen. This work describes a cultural enrichment method coupled with an H. pylori‐specific PCR to identify these bacteria in water. While far from perfected at the present time, this represents an exciting new approach to studying the significance of water as a transmission mechanism for H. pylori. Evidence is presented that indicates culturable H. pylori bacteria were found using this enrichment/PCR method in a local groundwater source.  相似文献   

RECENT DEVELOPMENTS IN MULTIVARIATE CALIBRATION   总被引：1，自引：0，他引：1  

B.R.KOWALSKI M.B.SEASHOLTZ Laboratory for Chemometrics  BG-  University of Washington  Seattle  WA  U.S.A. 《地理学报(英文版)》1991,(3)

With the goal of understanding global chemical processes,environmental chemists have some of the mostcomplex sample analysis problems.Multivariate calibration is a tool that can be applied successfully inmany situations where traditional univariate analyses cannot.The purpose of this paper is to reviewmultivariate calibration,with an emphasis being placed on the developments in recent years.The inverseand classical models are discussed briefly,with the main emphasis on the biased calibration methods.Principal component regression(PCR)and partial least squares(PLS)are discussed,along with methodsfor quantitative and qualitative validation of the calibration models.Non-linear PCR,non-linear PLSand locally weighted regression are presented as calibration methods for non-linear data.Finally,calibration techniques using a matrix of data per sample(second-order calibration)are discussed briefly.  相似文献   

固定标本的DNA提取及PCR扩增   总被引：4，自引：1，他引：3  

高天翔  张秀梅  曾晓起  渡边精一 《中国海洋大学学报(自然科学版)》2000,30(2):244-248

报道福尔马林、酒精固定的日本绒螯蟹标本的 DNA提取及 PCR扩增 ,并与冷冻、煮沸标本进行了比较。结果表明 ,福尔马林固定标本与酒精固定标本一样可以提取 DNA,加入的蛋白酶 K量越多 ,DNA的回收率越高。以提取的 DNA为模板 ,进行了线粒体 DNA12 S r RNA和 D- loop基因片段的 PCR扩增 ,经琼脂糖凝胶电泳 PCR扩增产物后均得到了与期待的碱基长度相一致的清晰谱带。  相似文献