共查询到20条相似文献，搜索用时 62 毫秒
1.
笔者初步研究了企鹅珍珠贝（Pteria penguin）组织蛋白酶D（cathepsin D,CTSD）的基因克隆和功能,通过同源克隆方法和cDNA末端快速扩增技术（RACE）获得了企鹅珍珠贝组织蛋白酶D基因（命名为pgCTSD）。该基因cDNA全长1 767 bp,其中5＇UTR为38 bp,3＇UTR为553 bp,ORF为1 176 bp,编码392个氨基酸,包括信号肽（Met1-Ala18）、前体域（Leu19-Lys47）和成熟域（Tyr48-Ser392）三部分,分子量为42.3 kDa,等电点为8.04。pgCTSD氨基酸序列与大珠母贝（Pinctada maxima）pmCTSD的相似性最高（79%）,与其他物种的相似性为59%～75%。荧光定量分析表明,空白对照组中pgCTSD mRNA在闭壳肌、性腺、肝胰脏、外套膜和鳃组织中都有表达,且在闭壳肌中表达量最少,肝胰脏中最高。与试验对照组相比,脂多糖（LPS）刺激6 h后性腺和肝胰脏显著下降,闭壳肌的表达量虽不大但增加显著,外套膜和鳃组织变化不显著;哈维弧菌（Vibrio harveyi）刺激6 h后肝胰腺和外套膜显著下降,闭壳肌和鳃显著上升,性腺无显著变化。肝胰腺中pgCTSD对LPS和弧菌刺激的应答反应表明pgCTSD可能参与了免疫反应。 相似文献
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Masato KINOSHITA Taijiro YABE Mitsutoshi KUBOTA Kazuharu TAKEUCHI Satoshi KUBOTA Haruhiko TOYOHARA Morihiko SAKAGUCHI 《Fisheries Science》2002,68(3):618-626
SUMMARY: Toughness is one of the most important elements that define the commercial value of the raw meat of fish. Degradation of the extracellular matrix is thought to be a cause of postmortem tenderization of fish meat. A previous study has suggested that this tenderization is caused mainly by metalloproteinases. The present study seeks to identify the proteinase(s) involved in tenderization; hence, cloned cDNA of two gelatinases from Japanese flounder, which showed high homology with mammalian matrix metalloproteinase (MMP)-2 and MMP-9, were designated as jfMMP-2 and jfMMP-9 , respectively. Northern blot analysis revealed that jfMMP-2 mRNA was expressed almost ubiquitously in adult tissues including the brain, muscle, gill, heart, gut, kidney, spleen, testis, and ovary. In contrast, the expression of jfMMP-9 mRNA was observed in those tissues which were abundant in blood cells, such as kidney, spleen, heart, and gill. Both recombinant proteins (jfMMP-2 and jfMMP-9) produced with the COS-7 cell system exhibited gelatin-degrading activity that was sensitive to 1,10-phenanthroline, a typical metalloproteinase inhibitor. 相似文献
3.
基质金属蛋白酶(matrix metalloproteinases,MMP)是一种能够降解细胞外基质的蛋白水解酶。MMP-17是一种膜型基质金属蛋白酶,通过糖基磷脂酰肌醇连接于细胞表面,参与调控有机体的内环境稳定、宿主防御等多种生理过程。为研究MMP-17在马氏珠母贝免疫反应中的作用,实验运用RACE技术,克隆得到马氏珠母贝MMP-17(Pinctada martensii MMP,Pm-MMP-17)基因cDNA 全长序列,并对其序列特征及功能进行初步分析。结果显示,Pm-MMP-17基因cDNA全长2 794 bp,开放阅读框(ORF)为1 923 bp,编码640个氨基酸,5'UTR长156 bp,3'UTR长715 bp,分子量约为73.11 ku,等电点为8.98;多序列比对和系统进化树分析表明,Pm-MMP-17与其他物种的MMP具有较高的保守性,与长牡蛎的MMP-17相似性高达82%;功能结构域分析表明,Pm-MMP-17有5个高度保守的结构区域:N-末端的信号肽、前导区、催化区、铰链区和C-末端的类血红素结合蛋白区;荧光定量数据分析表明,Pm-MMP-17基因在马氏珠母贝的闭壳肌、珍珠囊、足、外套膜、血淋巴、肝胰腺、性腺、鳃等8个组织中均有表达,在血液中的表达量最高,闭壳肌和鳃次之;脂多糖(LPS)刺激后,Pm-MMP-17基因表达水平上调,12 h后达到最大值,之后又逐渐下调并恢复到正常水平。研究表明,Pm-MMP-17基因可能在马氏珠母贝的免疫反应中起着重要作用。 相似文献
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Kazuharu TAKEUCHI Satoshi KUBOTA Masato KINOSHITA Haruhiko TOYOHARA Morihiko SAKAGUCHI 《Fisheries Science》2002,68(3):610-617
ABSTRACT: We have cloned a cDNA encoding the MMP-9 from a carp epidermal cell (EPC) cDNA library. The clone contains a 2025-base pair (bp) open reading frame encoding a protein of 674 amino acids. The deduced amino acid sequence shares 68% and 69% identity with medaka and Japanese flounder MMP-9. The hinge domain of the carp MMP-9, like those of the other non-mammalian species, lacks a type V collagen-like region that is typical of mammalian MMP-9. Gelatin zymography and immunoblot analysis of conditioned media of EPC cells and cDNA-transfected COS-7 cells detected a 76-kDa gelatinase. The apparent molecular mass of the carp zymogen is much smaller than those of its mammalian counterparts while almost identical with that of chicken 75-kDa gelatinase B-like enzyme. Although hypo-osmotic stress induced the elevation of MMP-9 mRNA level in EPC cells, no significant change in the protein in conditioned medium was detected during hypo-osmotic stress. Northern blot analysis detected a large amount of MMP-9 mRNA in carp kidney and spleen, suggesting the high expression of MMP-9 in blood cells, neutrophils, and macrophages. The smaller amount of MMP-9 mRNA was detected in gill, heart, fin, and eye, whereas none of the mRNA was detected in the hepatopancreas, intestine, brain, muscle, and skin. 相似文献
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The Pacific oyster Crassostrea gigas is a sessile bivalve that inhabits the intertidal zone and therefore frequently exposed to air during the tidal cycle. It
is highly adaptive to hypoxic conditions. We have studied the physiological state of oysters during long-term exposure to
air. The oysters became hypoxic when exposed to air or hypoxic seawater. The 50% lethal time of oysters exposed to air at
4, 15 and 20°C was 47.8, 15.9 and 12.2 days, respectively. The hemolymph pH decreased by day 3; however, it showed a slight
increase by day 5 at both 4 and 20°C. The adenylate energy charge (AEC) values decreased rapidly on the first day of air exposure
in the adductor muscle, mantle, gill and body trunk, and these decreases were accompanied by decreases in ATP concentrations
and increases in AMP concentrations. The AEC values in all of the tissues had fallen to below 30% by day 50 of air exposure
at 4°C. These data suggest that the energy state of oysters deteriorates rapidly with air exposure. Consequently, AEC values
may be useful indices of the physiological state of the oyster during long-term exposure to air. 相似文献
6.
cDNA cloning of Japanese oyster stress protein homologous to the mammalian 78-kDa glucose regulated protein and its induction by heatshock 总被引：1，自引：0，他引：1
Yoshihiro Yokoyama Hisashi Hashimoto Satoshi Kubota Akira Kuriyama Yoichi Ogura Shoshi Mizuta Reiji Yoshinaka Haruhiko Toyohara 《Fisheries Science》2006,72(2):402-409
ABSTRACT: The 78-kDa glucose regulated protein (GRP78), a member of stress proteins, was cloned from a cDNA library of Japanese oyster Crassostrea gigas . The analysis on Japanese oyster GRP78 clone of approximately 2.6 kb revealed that the entire open reading frame was 1983 bp long and encoded 661 amino acid residues. At the DNA sequence level, the coding region of Japanese oyster GRP78 gene was 72, 62, and 62% identical to those of chicken GRP78, Japanese flounder HSP70, and Japanese flounder HSC71 genes, respectively. Deduced amino acid sequence of Japanese oyster GRP78 was 84, 62, and 62% identical to those of chicken GRP78, Japanese flounder HSP70, and Japanese flounder HSC71, respectively. Japanese oyster GRP78 contained an 18-residue sequence at the N-terminus that exhibits characteristics of a cleavable signal sequence. It also contained an ATPase domain, and a peptide-binding domain in addition to a Lys-Asp-Glu-Leu (KDEL) peptide motif that is involved in determining endoplasmic reticulum localization. Northern blot analysis showed that GRP78 mRNA was induced with heatshock treatment in the oyster tissues. 相似文献
7.
Yan Li Jian G. Qin Xiaoxu Li Kirsten Benkendorff 《Aquaculture (Amsterdam, Netherlands)》2009,286(1-2):64-71
This paper investigates the temporal responses of 2-year old Crassostrea gigas to environmental changes in Stansbury, South Australia from September 2005 to October 2006. A total of 360 oysters were grown in one-line baskets on the farm using six replicates that were sampled monthly. A range of environmental parameters were assessed and correlated against biological indicators for oyster condition, metabolism and antimicrobial activity. Food availability by chlorophyll a, was low throughout the study period (0.5–1.5 µg L? 1) and was significantly correlated to phosphorus concentrations. The condition index and shell weight of oysters significantly increased over the year, with the condition index dropping after spawning but then recovering within one month. Significant temporal variation in energy storage and utilization were observed in different tissues over the year. Glycogen in the mantle tissue was influenced by reproduction and correlated to chlorophyll a levels, but not in the gill or adductor muscle. The mantle glycogen and gill protein reached the lowest level in February when spawning occurred and presented evidence for seasonal variation in oyster metabolic activity. However, mantle and adductor muscle proteins did not drop after spawning indicating that these proteins contribute little to gametogenesis. Hemolymph protein was negatively correlated to water temperature and chlorophyll a, reaching the lowest level during summer. Hemolymph antibacterial activity significantly decreased after spawning, implying that the period of post-spawning is critical for oyster health. This study revealed trade-offs in the energy budget between immune resistance, growth, and reproduction. The results indicate that in a lean water environment, spawning events significantly regulate metabolic and immune capacities of oysters and a second year of rearing increased meat and shell weight but not the shell length. These findings are applicable to the management and development of oyster aquaculture within temperate southern hemisphere. 相似文献
8.
虾夷扇贝组织中微量元素的分布特性 总被引：1，自引：0，他引：1
研究了虾夷扇贝闭壳肌、外套膜、内脏团、瓣鳃和性腺中Cu、Fe、Mn、Zn、Pb和Cd等元素的分布特性。试验结果表明,内脏团中的Cu显著(P<0.05)高于其他各组织,其他各组织间差异不显著(P>0.05);闭壳肌中Zn的含量显著(P<0.05)高于内脏团、外套膜、瓣鳃和性腺中的含量;Fe含量内脏团和瓣鳃中显著(P<0.05)高于性腺、闭壳肌和外套膜;各组织中Mn的分布特性为瓣鳃>闭壳肌>性腺>内脏团>外套膜;Pb的分布特性为内脏团>闭壳肌>性腺>外套膜>瓣鳃;内脏团中的Cd占全贝总量的67%,显著高于其他组织。因此,约占全贝质量10%的内脏团蓄积了较高含量的Cu、Fe、Pb和Cd,尤其是Cu和Cd(分别约占全贝蓄积总量的71%和67%),食用时去掉内脏团,可保证虾夷扇贝的食用安全和较高的食用及营养价值。 相似文献
9.
基质金属蛋白酶(MMPs)是一种能够降解细胞外基质的蛋白水解酶类。为研究MMPs在仿刺参免疫防御中的作用,本实验采用RACE技术克隆了仿刺参基质金属蛋白酶16基因(Aj-MMP-16)的cDNA全长序列,并对其序列特征和功能进行了初步分析;采用实时荧光定量PCR(qRT-PCR)方法,分别分析了Aj-MMP-16基因在仿刺参不同组织、不同"化皮"体壁组织以及病原菌刺激后体腔细胞中的表达情况。结果显示,Aj-MMP-16基因的cDNA全长为2 976 bp,包括一个342 bp的5′非编码区,一个963 bp的3′非编码区;开放阅读框(ORF)为1 671 bp,编码557个氨基酸,预测蛋白分子量为63.11 ku,等电点为4.79。Aj-MMP-16具有典型的MMPs家族蛋白结构:N-端前肽区、铰链区、催化区、C-端类血红素结合区和跨膜区。Aj-MMP-16与其他物种的MMPs具有一定的相似性,与紫色球海胆的MMP-16相似性最高。Aj-MMP-16基因mRNA在仿刺参各组织中均有表达,表达量由高到低为呼吸树、肠、体腔细胞、管足、肌肉、体壁;在"化皮病"不同阶段,AjMMP-16基因mRNA在"化皮"体壁组织中的表达量显著高于正常体壁组织;灿烂弧菌和蜡样芽孢杆菌刺激后,体腔细胞中Aj-MMP-16基因mRNA表达量显著升高。Aj-MMP-16基因可能在仿刺参内脏再生、炎症发生以及免疫应答中起着重要的作用。 相似文献
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11.
Masatomi Hosoi Seiya Sugihara Kouta Kato Shoshi Mizuta Yoshihiro Yokoyama 《Fisheries Science》2014,80(4):819-825
The effects of immersion salinity on the food properties [water content, salinity, and free amino acid (FAA) content] of shucked oysters were analyzed. Results of a laboratory immersion experiment suggested that the molluscous parts (other than the adductor muscle) swelled in lower salinity and shrank in higher salinity. Higher FAA content was observed in oysters immersed in higher-salinity water. In the adductor muscle, water content increased and FAA content decreased markedly following immersion, regardless of salinity, probably because of intake of immersion fluid and leakage of FAAs across the cut end of the adductor muscle. Immersion salinity ranged from 0.17 to 1.54 % in shucked oyster products on the retail market. Tissue salinity was strongly correlated with immersion salinity (r = 0.904), and tissue water content was correlated negatively with immersion salinity (r = ?0.668). In addition, total FAA and taurine content of oysters were correlated with immersion salinity (r = 0.629 and 0.865, respectively). These results clearly indicate that immersion salinity is an important factor affecting the food components of shucked oysters. 相似文献
12.
紫贻贝SOD和POD同工酶的组织特异性研究 总被引：5，自引：0，他引：5
用聚丙烯酰胺垂直电泳对紫贻贝的鳃、性腺、外套膜、足、闭壳肌等5种组织的超氧化物歧化酶(SOD)和过氧化物酶(POD)进行了检测分析。结果表明,SOD在各组织器官中均表达,共表现为5条酶带,酶带a和b在上述五种组织中均出现,酶带c在除鳃以外的其他4个组织中有表达,酶带d则在除外套膜以外的4个组织中表达,酶带e仅在性腺中存在,所有上述酶带在性腺和鳃中活性相对较强;POD在不同组织中表达各不相同,酶带a仅在外套膜中存在,酶带b存在于除外套膜外的4个组织中,酶带c在性腺、闭壳肌和外套膜中表达。从检测结果来看,SOD和POD在紫贻贝体内的表达有一定的组织特异性,这种特异性不仅与各组织的不同生理分工相关,而且也与机体对自身基因表达进行调控相联系。 相似文献
13.
ABSTRACT: Four toxins, okadaic acid (OA), dinophysistoxin-1 (DTX1), pectenotoxin-6 (PTX6), and yessotoxin (YTX), all associated with diarrhetic shellfish poisoning (DSP), were administered via syringe to Scallops Patinopecten yessoensis and their distribution in the hepatopancreas, adductor muscle, and combined other tissues (mantle, gill, gonad) was analyzed by liquid chromatography-mass spectrometry. Toxins exclusively remained in the hepatopancreas irrespective of the injection site, adductor muscle or hepatopancreas. When injected into hepatopancreas, OA, DTX1, and YTX were metabolized to 7- O -palmitoylOA, 7- O -palmitoylDTX1 and 45-hydroxyyessotoxin (45OH-YTX), respectively. Such metabolic changes were insignificant when toxins were injected into the adductor muscle. The residual ratio for each toxin in the hepatopancreas was less than 20%. Mortalities of scallops treated with PTX6 were lower than those treated with other toxins. 相似文献
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为了解长牡蛎MITF基因的表达及其与壳色的关联,验证了长牡蛎中的4个MITF基因,对长牡蛎MITF氨基酸序列进行了序列分析和多序列比对,分析了长牡蛎幼体发育各时期的转录组,采用荧光定量PCR的方法研究长牡蛎各组织及黑壳、白壳牡蛎特定部位mRNA表达情况。4个MITF基因中有3个基因可能为假基因,有表达的长牡蛎MITF基因共编码448个氨基酸,为亲水性不稳定蛋白,含有N端结构域(MITFTFEBC3N superfamily)和高度保守的功能性结构域HLH结构域。转录组分析发现MITF在长牡蛎个体发育的各个时期均有表达,在稚贝期达到最高。组织表达结果显示MITF在外套膜中的表达水平显著高于其他组织。MITF在黑壳牡蛎闭壳肌中的表达量显著低于白壳牡蛎,而在黑壳牡蛎外套膜中的表达量高于白壳牡蛎,但不显著。在黑壳、白壳牡蛎外套膜边缘的表达量都显著高于内侧。研究表明,长牡蛎MITF基因可能在牡蛎壳形成早期就参与了黑色素的生成调控和个体的生长发育,可调控牡蛎酪氨酸酶Tyr2基因,参与外套膜和贝壳中黑色素的形成。本研究为进一步研究牡蛎壳色形成机制奠定了基础。 相似文献
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采用同源克隆和RT-PCR技术对合浦珠母贝(Pinctada fucata)热休克蛋白hsp70基因进行了克隆和表达分析。获得cDNA全长序列2 365 bp,其中3’非编码区域(UTR)为318 bp,5’UTR为88 bp,开放阅读框(ORF)为1 959 bp,编码652个氨基酸,分子量约为71.39 kD,理论等电点为5.22,并含有3个HSP70家族的签名序列IDLGTTYS、DLGGGTFD和EEVD。同源性分析表明,合浦珠母贝HSP70的氨基酸序列与太平洋牡蛎(Crassostrea gigas)等双壳贝类的相似性高达86%以上,基于氨基酸序列的聚类分析表明,合浦珠母贝与牡蛎属种类亲缘关系最近。高温、高盐刺激后,半定量RT-PCR检测发现hsp70基因的表达明显增加,高温刺激的表达量高于高盐刺激,高温刺激组不同组织的表达量由大到小依次为鳃、消化腺、外套膜、肌肉、性腺,高盐刺激组不同组织的表达量由大到小依次为鳃、外套膜、肌肉、消化腺、性腺,表明HSP70参与了机体对刺激的应答过程。该基因的克隆为进一步深入研究合浦珠母贝的抗逆机理及其遗传改良奠定了重要基础。 相似文献
17.
Qin Li Yaoguo Li Xiande Huang Maoxian He 《Journal of the World Aquaculture Society》2014,45(6):638-651
Myostatin (MSTN or growth differentiation factor‐8) is considered a negative regulator of muscle growth and development. In this study, we cloned and characterized the full‐length MSTN cDNA from Pinctada fucata, and named it as the Pf‐MSTN cDNA. The single nucleotide polymorphisms (SNPs) in Pf‐MSTN cDNA were then screened and genotyped. The full‐length Pf‐MSTN cDNA was 2644 bp, including an open reading frame of 1248 bp encoding 415 amino acids which contained typical structural characteristics shared by all members of the transforming growth factor‐β (TGF‐β) superfamily including an N‐terminal signal peptide, a propeptide domain, and a TGF‐β superfamily bioactive domain. The Pf‐MSTN mRNA was detected in all tested tissues, with the highest mRNA levels observed in the adductor muscle, indicating that Pf‐MSTN may play a major role in this tissue. Furthermore, by sequencing and alignment, 32 SNP loci were identified in Pf‐MSTN cDNA. Genotyping 50 individuals from a common breeding stock revealed that 21 of these 32 loci were polymorphic. The minor allele frequency was in the range of 0.0400–0.4800, and the polymorphism information content value varied from 0.0739 to 0.3750. The observed and expected heterozygosity ranged from 0.0200 to 1.0000 and from 0.0776 to 0.5051, respectively. These SNPs identified in Pf‐MSTN will be useful for future studies investigating their utility in marker‐assisted selection for P. fucata breeding. 相似文献
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采用逆转录聚合酶链式反应（RT－PCR）方法,从哲罗鲑（Hucho taimen）肝脏的总 RNA 中扩增出胰岛素样生长因子－I（IGF－I）的 cDNA 开放阅读框（Open reading frame, ORF）序列,运用软件对其进行生物信息学分析,并利用荧光实时定量 PCR 技术检测了哲罗鲑成鱼不同组织中 IGF－I mRNA 的表达情况。结果显示, IGF－I 基因的 cD－NA 开放阅读框为573 bp,编码190个氨基酸,蛋白质等电点为9．21,氨基酸结构由信号肽、 B、 C、 A、 D 结构域及 E 肽组成;氨基酸序列与其他鲑科鱼类具有较高的同源性,其中与北极红点鲑的 IGF－I 同源性最高（99．2％）;组织表达分析显示,哲罗鲑 IGF－I mRNA 在肝脏中表达量最高,在鳃、前肠中次之,在脑、头肾、脾、心、胃和肌肉等组织中的表达量较低。 相似文献
19.
为探讨干露胁迫对海产贝类基因组DNA甲基化的影响,应用荧光标记甲基化敏感扩增多态性(fluorescencelabeled methylation sensitive amplified polymorphism,F-MSAP)技术,比较了不同干露条件下(0 d、0.5 d、1 d、3 d、5 d、7 d、9 d和11 d)长牡蛎(Crassostrea gigas)基因组DNA甲基化的变化。结果表明,对照组(干露处理0 d)闭壳肌与鳃组织的总体甲基化水平分别为29.76%和29.82%;干露处理0.5 d、1 d、3 d、5 d、7 d、9 d和11 d的长牡蛎全基因组甲基化水平呈现先增高后降低的趋势,其中,闭壳肌组织的总体甲基化水平分别为36.59%、38.86%、43.02%、39.30%、51.13%、46.79%和35.06%,鳃组织总体甲基化水平分别为39.39%、42.13%、39.36%、43.54%、56.19%、38.57%和28.99%;干露处理7 d的长牡蛎甲基化水平明显高于其他时期(P0.05),11 d时甲基化水平基本恢复至初始状态。甲基化变异模式分析发现,闭壳肌与鳃组织DNA甲基化变异位点存在差异,甲基化升高位点变化程度较大(P0.05)。以上结果表明,长牡蛎通过改变DNA甲基化模式来应答干露胁迫,发生了不同程度的甲基化与去甲基化反应,DNA甲基化与长牡蛎的抗逆性状密切相关。 相似文献
20.
A full-length cDNA encoding the insulin-like growth factor binding protein-3 (IGFBP-3) was cloned from the liver of common carp (Cyprinus carpio) by RT-PCR. The IGFBP-3 cDNA sequence is 1,680 bp long and has an open reading frame of 882 bp encoding a predicted polypeptide of 293 amino acid residues. The deduced amino acid sequence contains a putative signal peptide of 25 amino acid residues resulting in a mature protein of 268 amino acids. A single band of approximate 1.9 kb was found in liver by Northern blot analysis. IGFBP-3 mRNA was observed in all regions of brain with high levels. In peripheral tissues, high levels of IGFBP-3 mRNA were found in retina, red muscle, liver, heart, posterior intestine, spleen, and testis. Relatively lower levels were found in white muscle, kidney, thymus gland, and ovary, while in head kidney, blood, skin, gill, middle intestine, and anterior intestine, the IGFBP-3 mRNA levels were much lower. IGFBP-3 mRNA was first detected in the blastula stage with significantly high level. The level sharply decreased in gastrula stage, and it became to increase in the following stages. During the reproductive cycle, the abundance of IGFBP-3 mRNA significantly decreased between the recrudescing stage and the matured stage in ovary, although in testis, IGFBP-3 mRNA expression level did not exhibit a significant change. The mRNA expression profiles in the present study imply that the IGFBP-3 may play important physiological functions in common carp development and reproduction. 相似文献