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1.
为了探讨三角帆蚌(Hyriopsis cumingii)在不同钙离子浓度养殖时,内脏团和外套膜细胞对环境中钙离子的吸收,以及细胞内钙离子浓度对碳酸酐酶的影响,本实验设定了5种钙离子的环境浓度(0 m M、0.5m M、1.25 m M、2 m M、3 m M),采用流式细胞仪测定内脏团和外套膜组织细胞内钙离子含量,用荧光定量PCR检测α-碳酸酐酶基因(HcCA)表达,并用乙酸对硝基苯酯的水解间接测定碳酸酐酶活性,以期能阐明环境中钙浓度对组织细胞钙含量的影响,和组织细胞内不同钙含量与碳酸酐酶基因表达和酶活性之间的关系。研究结果表明,在相同环境的钙离子浓度下,活体三角帆蚌的内脏团细胞中钙含量显著高于外套膜细胞(P0.05),并且表明从0 m M到2 m M浓度,内脏团和外套膜细胞内的钙含量显著上升(P0.05),在3 m M浓度时出现下降(P0.05)。同时,细胞内Ca~(2+)含量较低时,HcCA在内脏团和外套膜中的表达量均显著低于细胞内Ca~(2+)含量较高组(P0.05),但细胞内Ca~(2+)荧光强度为8×104时达到饱和且开始下降,而HcCA的表达在细胞内Ca~(2+)荧光强度为6×104已经最高,并开始下降。环境钙离子浓度在0 m M、0.5 m M时碳酸酐酶活性显著高于1.25 m M、2 m M、3 m M浓度组(P0.05),而且1.25 m M、2 m M、3 m M浓度组间无显著差异(P0.05),由此发现碳酸酐酶在0.5 m M钙离子浓度中活性较高但表达量却较低,而在1.25 m M和2 m M浓度中表达量高但活性较低,并且碳酸酐酶外套膜细胞酶活性在各浓度均显著高于内脏团细胞(P0.05)。本研究为三角帆蚌水体最适养殖钙浓度提供了重要依据。  相似文献   

2.
三角帆蚌外套膜表达的免疫相关基因筛选   总被引:1,自引:0,他引:1  
利用三角帆蚌(Hyriopsis cumingii)外套膜cDNA文库EST序列,通过BLAST分析注释基因功能,发现35个与免疫防御功能相关的基因。根据其功能,这些免疫相关基因可划分为7类,即细胞免疫过程(2个)、蛋白酶和蛋白酶调节子(9个)、压力蛋白(7个)、抗菌肽(5个)、溶酶体酶(2个)、粘着蛋白(3个)及细胞凋亡和细胞周期调控相关基因(7个)。免疫相关基因在三角帆蚌外套膜中的广泛表达表明外套膜是重要的免疫组织。该研究为三角帆蚌外套膜分子免疫机理研究和抗病育种工作提供了基础性资料。  相似文献   

3.
溶血磷脂酰胆碱酰基转移酶 1(LPCAT1)是一种重要的脂质代谢酶。为明确三角帆蚌(Hyriopsis cumingii) HcLPCAT1 基因在类胡萝卜素代谢中的功能, 探究该基因与三角帆蚌壳色的相关性, 本研究采用 RACE 技术克隆获得 HcLPCAT1 基因 cDNA 全长 1675 bp, ORF 区 1296 bp 编码 431 个氨基酸; 实时荧光定量分析发现 HcLPCAT1 基因在紫色三角帆蚌各组织中表达量均高于白色三角帆蚌相应组织, 且在肝胰腺、外套膜中的表达量差异显著 (P<0.05); 原位杂交检测到的阳性信号主要定位在外套膜的外褶、背膜区、腹膜区, 外褶与中褶连接处以及部分中褶; 紫色三角帆蚌补充投喂 β-胡萝卜素后, HcLPCAT1 基因在肝胰腺、中央膜、边缘膜各组织中表达量极显著上调 (P<0.01), 同时相应组织中总类胡萝卜素含量(TCC)极显著升高(P<0.01)。采用直接测序法鉴定出三角帆蚌 HcLPCAT1 基因 5 个 SNP 位点的基因型与内壳色存在显著相关性(P<0.05), 单倍型分析发现 H1、H2 两种单倍型为紫色三角帆蚌优势单倍型, H3、H5、H6 三种单倍型为白色三角帆蚌优势单倍型。本研究鉴定的 HcLPCAT1 基因可为解析三角帆蚌类胡萝卜素代谢和壳色形成的机制提供研究基础, 筛选的与内壳色相关 SNP 及单倍型可用于分子辅助育种。  相似文献   

4.
观察了不同pH值对三角帆蚌珍珠形成的影响.结果显示,在pH 5、6、7、8、9时,三角帆蚌鳃组织中的钙含量最高,远远高于外套膜和珍珠囊.在pH为5~8时,随着pH值的升高,鳃和外套膜组织中的总钙量逐渐增加.在pH 6时,珍珠囊和珍珠的长、短轴直径最大,分别为(8.027±0.759) mm、(6.817±0.772) mm、(7.187±0.850) mm、(6.157±0.810) mm,质量最大,分别为(0.494±0.153)g、(0.419±0.140)g,与其他pH值水体的三角帆蚌相比,差异显著(P<0.05).结果表明,pH 6的弱酸性水有利于三角帆蚌的生长发育,所形成的珍珠最大.  相似文献   

5.
为了发掘更多三角帆蚌具有EF-hand结构域的功能基因及其蛋白质,本研究运用RACE-PCR技术,克隆得到了三角帆蚌包含EF-hand结构域钙结合蛋白1基因(EF-hand calcium-binding domain-containing protein 1,EFCB1)的cDNA全长并进行了生物信息学分析;通过real-time Q-PCR技术,分析了EFCB1基因在三角帆蚌10个组织,以及内脏团、外套膜插核后不同时间点的时空表达特点。结果表明三角帆蚌EFCB1基因cDNA序列全长981 bp,ORF为531 bp,编码176个氨基酸残基,5'-UTR 239 bp和3'-UTR 211 bp。EFCB1分子式为C877H1348N238O270S10,分子量约19.9 ku,等电点为4.70,不稳定系数为62.65,属亲水蛋白。其序列无信号肽序列,存在1个跨膜区域和2个EF-hand结构域,EF-hand模块分别为DLNDDKLISPEE(98-109)和DTNGDDKLDGEE(129-140)。荧光定量结果显示三角帆蚌EFCB1基因在各组织中均有表达,其中在肠和鳃中表达量最高(P<0.05),外套膜中表达量显著高于内脏团(P<0.05)。EFCB1基因在插核后不同时期的外套膜和内脏团育珠部位组织中表达具有显著差异(P<0.05),在外套膜中的表达量均显著高于内脏团(P<0.05),在插核后第20 天时表达量显著高于各时期(P<0.05)。研究表明,EFCB1在三角帆蚌Ca2+的吸收过程中发挥调节作用,在珍珠囊形成过程中以及珍珠形成初期具有重要功能。  相似文献   

6.
珍珠与珍珠蚌内壳珍珠层具有相似的形成机制,已发现珍珠颜色与供片蚌内壳色显著相关。该研究以紫色、金色、白色3种色系三角帆蚌(Hyriopsis cumingii)为对象,设置β-胡萝卜素补充实验组和对照组,养殖90d后比较分析了不同色系三角帆蚌内壳色、组织总类胡萝卜素含量(TCC)及生长变化。结果表明,实验组紫色三角帆蚌内壳色较对照组色差值(dE)提高了21.48%(P0.05),明度值(L~*)降低了15.72%(P0.05),色度值(a~*)从0.48升至2.67 (P0.05),色度值(b~*)未见显著变化(P0.05);实验组金色三角帆蚌内壳色较对照组a~*从0.07升至1.52 (P0.05),b~*从1.37升至4.43 (P0.05),dE和L~*未见显著变化(P0.05);实验组白色三角帆蚌内壳色各参数较对照组均未见显著变化(P0.05)。3种色系三角帆蚌实验组肝胰腺TCC均大于对照组(P0.05);紫色和金色实验组外套膜TCC较对照组分别提高了55.29%和39.69%(P0.05),白色实验组较对照组未见显著变化(P0.05)。实验组3种色系三角帆蚌各生长性状均大于对照组(P0.05)。研究结果证实补充β-胡萝卜素可提升三角帆蚌的内壳色和促进生长,为珍珠养殖技术优化提供理论依据。  相似文献   

7.
三角帆蚌钙网蛋白基因cDNA的分子特征与表达分析   总被引:2,自引:2,他引:0  
为研究淡水珍珠形成相关基因及调控机理,以三角帆蚌为研究对象,借助cDNA末端快速扩增(RACE)技术获得了三角帆蚌的钙网蛋白基因(calreticulin,HcCRT) cDNA序列,该序列长1 437 bp,包含231 bp的5'非编码区(untranslated region,UTR)和615 bp 3'UTR以及591bp的开放阅读框(ORF),共编码196个氨基酸残基,包含一段由21个氨基酸组成的信号肽和一段由175个氨基酸的成熟肽,分子量约为22.4 ku,理论等电点5.01.氨基酸序列分析表明,该序列不存在跨膜结构,疏水性分析显示该蛋白整体为亲水性蛋白.氨基酸列比对分析显示,HcCRT具有保守的钙网蛋白家族结构,具有2个保守的钙网蛋白家族标签序列:KHEQNIDCGGGYLKVF和IMFGPDICG,与其他已知物种的CRT具有较高保守性,其中与斑马鱼的相似度为77%,与长牡蛎和马氏珠母贝的相似度为70%.对三角帆蚌HcCRT蛋白序列的二级结构和三级结构进行预测分析,显示该蛋白同时含螺旋和折叠.实时荧光定量PCR分析结果显示,HcCRT在外套膜、血液、鳃、斧足、肝脏、肾脏、肠和闭壳肌等8个组织中均有表达,其中在外套膜中表达量最高,血液次之,在其他组织的表达量极少.初步推测HcCRT参与了三角帆蚌珍珠形成的生物矿化过程.  相似文献   

8.
清道夫受体(SR)是一类对化学修饰的脂蛋白具有很强结合活性的糖蛋白家族。本研究通过RACE方法克隆得到三角帆蚌hcSRCR1基因cDNA序列,该序列全长1 000 bp,其中开放阅读框819 bp,编码272个氨基酸,预测分子量为28.16 ku,理论等电点为5.55;预测含有2个SRCR结构域和6个保守的半胱氨酸残基。qRT-PCR和Western blot结果显示,hcSRCR1 mRNA和蛋白表达模式基本相同,均在三角帆蚌外套膜中表达量最高,在其他组织中的表达量普遍较低,且在紫色选育系外套膜组织中的表达量显著高于白色选育系。外套膜原位杂交结果显示,hcSRCR1基因主要在外套膜外褶的内、外上皮细胞层以及腹膜处的上皮细胞层中表达。研究表明,三角帆蚌hcSRCR1基因与贝壳珍珠质颜色形成具有一定相关性,可为进一步研究该基因在珍珠颜色形成过程中的调控机理提供基础资料。  相似文献   

9.
众所周知,优质珍珠大多产于三角帆蚌,但在三角帆蚌易发病,目前尚未得到很好控制之前,采用三角帆蚌育珠风险较大。褶纹冠蚌生长快,抗病力强,只要采取一定的措施,同样也能生产出优质珍珠。现将我们的育珠经验介绍如下:一、亲蚌选择和小蚌培育褶纹冠蚌以1一'2齿令时期生长最旺盛,之后生长逐渐缓慢据吴江县东太湖八都水产场对1一2龄的褶纹冠蚌做手术蚌后在同等条件下的产珠比较(见表1)叮证明:l龄蚌优于'2龄蚌然而作为手术蚌,还要求外套膜有一定的厚度,插片操作方便,因此蚌体必须大于8厘米如何把1龄蚌当年培养到8一10厘米是首要前提_试验证明,只要措施得当,大部分的蚌是能够达到这一要求  相似文献   

10.
对三角帆蚌(Hyriopsls cumingii Lea)进行内脏团插核手术,并对术后机体免疫相关因子基因alpha-2巨球蛋白(α2M)、酶酸性磷酸酶(ACP)和超氧化物歧化酶(SOD)的变化情况进行研究,旨在为三角帆蚌内脏团育珠实践提供理论依据.实验选取100只健康三角帆蚌分为2组(各50只,每组各设5个重复,每重复各10只蚌),一组在蚌体内脏团进行插核手术(实验组),一组未经处理(对照组),分别饲养于相同条件恒温(24℃)淡水缸内.两组分别于插核后1天、2天、3天、5天及10天采集淋巴血,通过RT-PCR及酶活测定法研究α2M基因表达和ACP、SOD在术后活性的变化.结果发现,α2M基因在手术后表达量增加,在插核后第3天和第5天与对照组差异显著(P<0.05);实验组血清和血细胞中ACP活性均显著高于对照组;实验组血清中SOD活性在第3天、5天、10天高于对照组(P<0.05);实验组血细胞中SOD的活性低于对照组.本研究表明,内脏团插核手术后三角帆蚌机体的免疫防御调节增强,血液中免疫相关基因α2M的表达水平和免疫相关酶ACP和SOD活性均有明显变化.  相似文献   

11.
The purpose of the present study is to investigate the gonadosomatic index (GSI %), the absolute and relative fecundity of one-year-old pike weighing over 400 g and at the same time to study the dependence between the egg size and the size, behavior and vitality of the free embryos obtained from one- and two-year-old spawners. The study involved two weight groups of females, differing in age and body weight and length—one-year-old (W = 514 g, SL = 36.1 cm) and two-year-old matured pike (W = 1454 g, SL = 49.3 cm). Ovary samples were fixed and egg follicles containing maturing oocytes counted and weight. The weight and length of the free embryos from semi-artificial spawning were measured. The results showed that, when raising this species under farmed conditions, more than 40% (in rarer cases 90%) of one-year-old pike females reached over 400 g and 35 cm (SL) and reached puberty. Absolute fecundity of 15,030 follicles (30 follicles per gram body weight) was observed; the GSI was nearly 15% and the follicle weight reached 3.7 mg. This data differs significantly from that obtained from the larger two-year-old fishes: absolute fecundity 41,363 follicles (28 follicles per gram body weight), GSI nearly 20%, follicle weight 5.8 mg. Results showed that the different follicle size determines the free-embryo size. A positive linear correlation was found between the egg follicle weight and the free-embryos weight (r = 0.7143). The free embryos obtain from the one- and two-year-old spawners differed significantly both in terms of their weight (7.13 mg against 10.61 mg) and total length (0.81 cm against 0.97 cm), the differences being 1.5- and 1.2-fold, respectively.  相似文献   

12.
13.

DMEM为基础培养基, 通过优化改良培养基及缓冲液的配方, 对三角帆蚌(Hyriopsis cumingii Lea)外套膜进行细胞培养, 并且以显微观测、RNA/DNA的比值作为该细胞增殖的评价指标, 分别对体外培养细胞的迁出时间、速度、数量以及增殖活力进行测定。分别取体外培养第24108120 小时的细胞进行Hoechst DNA荧光标记, 然后将3组标记细胞植入三角帆蚌外套膜中在蚌体内培养, 在植入后第2472120168216 小时运用RNA/DNA指标检测活体培养的细胞增殖活力。结果表明, 经过优化后的缓冲液和培养基更有利于细胞从外套膜组织中迁出, 迁出速度和细胞数量显著增加(P<0.05), 且细胞的活力随着培养时间的延长而逐渐增大, 培养至108 h, 细胞活力达到最大, RNA/DNA比值为 24.53, 108 h后显著下降(P<0.05), 显微观测的细胞生长状况与RNA/ DNA指标测定相吻合。对体外培养的细胞植入蚌体外套膜中再进行培养时发现, 对体外培养活力较好的细胞, 活体内环境可增大细胞的活力, RNA/DNA比值最高达到25.45, 但在活体培养168 h 后活力显著下降(P<0.05); 而对体外培养活力较差的细胞, 活体内环境对细胞活力影响不显著(P>0.05)。本研究旨在为三角帆蚌外套膜细胞增殖的深入研究及建株提供基础性资料。

  相似文献   

14.
三角帆蚌HcCUBDC基因cDNA的全长克隆与表达分析   总被引:1,自引:1,他引:0  
利用cDNA末端快速扩增(Rapid Amplification of cDNA Ends,RACE)方法,获得了三角帆蚌HcCUBDC基因的全长cDNA序列,共5158bp,开放阅读框(ORF)1920bp,编码639个氨基酸,5′端非编码区576bp,3′端非编码区2662bp,GeneBank登陆号为KP067952。生物信息学分析表明,三角帆蚌HcCUBDC蛋白含有一个由19个氨基酸组成的信号肽和4个CUB结构域,但未能比对上已知的蛋白。经荧光定量PCR检测,HcCUBDC基因在紫色和白色蚌前端缘膜、后端缘膜、中央膜、鳃、斧足、肝胰腺、肠和肾8个组织中均有表达,并且都是肝胰腺表达量最低。在紫色蚌中,后端缘膜表达量极显著高于前端缘膜;在白色蚌中,前端和后端缘膜之间表达差异不显著。HcCUBDC基因在紫色蚌后端缘膜中的表达量极显著高于白色蚌,在紫色和白色蚌前端缘膜中的表达量差异不显著。外套膜原位杂交结果显示,HcCUBDC基因主要在缘膜外上皮细胞中表达。研究表明,HcCUBDC基因在三角帆蚌内壳色形成中发挥作用,可为进一步深入研究该基因在珍珠颜色形成过程中的功能及其调控机理提供基础资料。  相似文献   

15.
Pinctada fucata martensii is an ideal animal for study of biomineralization. Although dozens of genes have been identified, the molecular mechanism of biomineralization remains still unclear. The purpose here was to discover the expression profiles of nine biomineralization genes in related tissues: mantle edge (ME), mantle centre (MC) and pearl sac (PS), and to explore the relationships between expression level and nacre thickness. The expression levels of seven genes (ACCBP, aspein, CaM, EFCBP, KRMP, nacrein and prismalin‐14) showed no significant difference between ME and MC, but were significantly higher than those in PS. The expression level of N19 was the same in ME and PS, significantly higher than that in MC. However, msi60 in ME, MC and PS showed no significant differences in expression level. In addition, the correlations of expression levels between ME, MC and PS were highly significant, indicating that there are some similar function and mechanism among the three tissues. Also the nine genes had significant correlation of expression levels between one another. Furthermore, no other genes from oysters which produced pearls of different grades showed significant difference in the expression level, except CaM in MC, which had significantly higher expression level in the medium pearl grade than in the plain nucleus grade. Moreover, the expression level of msi60 in MC of host oyster was significantly correlated with produced pearl layer thickness, suggesting that msi60 might play an important role in pearl formation and host oyster may also participate in the development of pearl.  相似文献   

16.
Growth and pearl production were compared for males and females in the freshwater mussel, Hyriopis cumingii, from a full-sib family. The results indicated that there was no significant difference (p > 0.05) on individual weight between male and female mussels of 1- or 2-year-old, while significant difference (p < 0.01) lay among 3- or 4-year-old mussels with male greater than female. The average shell width of the male mussels was less than that of the female individual (p < 0.05). 1- and 2-year-old males and females did not differ significantly with respect to the total weights, grain weights, or grain sizes of the pearls they produced, but these three parameters were all significantly greater in 3- and 4-year-old males (p < 0.05). The round pearl percent were similar between male and females at ages 1, 2, 3 and 4 years. Male and female mussels were separately and mixed cultured in enclosures, respectively. The rates of growth in shell width and body weight of females separated from males were 3.42 and 4.16 %, respectively, higher than in females mixed with males (p < 0.05). The total pearl weight per mussel, the average weight per pearl, and the average pearl size of females separated from males were 6.61, 7.10, and 3.59 %, respectively, greater than in females mixed with males (p < 0.05). There was no significant difference in the growth rate or pearl yield of males cultured with or without females (p > 0.05). Under traditional culture methods, male mussels have a better pearl performance, and artificial separation of females from males can improve the growth and pearl production of female mussels.  相似文献   

17.
马氏珠母贝外套膜不同区域基因组DNA甲基化MSAP分析   总被引:1,自引:0,他引:1  
通过甲基化敏感多态性扩增(methylation-sensitive amplification polymorphism,MSAP)技术,检测马氏珠母贝(Pinctada martensii)外套膜的边缘膜区(mantle edge,Me)、套膜区(mantle pallial,Mp)和中央膜区(mantle central,Mc)的基因组DNA甲基化修饰水平;回收具特异性的清晰甲基化修饰片段进行测序、比对分析并筛选基因;利用荧光定量PCR对筛选基因进行定量分析。结果显示:(1)利用15对引物进行扩增实验,平均每个个体外套膜3个区域能够产生(1163.25±124.34)条清晰可辨的条带,其中Me、Mp和Mc分别得到(401.00±40.37)条、(380.63±52.39)条和(381.63±53.57)条扩增条带,各组织甲基化总条数差异不显著(P0.05);Me、Mp和Mc的基因组甲基化水平分别为(17.07±2.19)%、(15.48±2.34)%和(19.61±2.88)%,Mc和Mp具有显著性差异(P0.05);组织间的基因组甲基化水平由高到低排列依次是McMeMp。(2)对特异性片段进行回收、测序后,经在线及本地Blast比对,得到8条存在甲基化修饰的基因序列,其中3条有同源序列,基因注释为40S核糖体蛋白SA(40S ribosomal protein SA)、iHog(interference hedgehog)、锌指蛋白(zinc finger protein castor)。iHog仅在中央膜上具有全甲基化修饰,且E值较低,为筛选的目的基因。(3)荧光定量结果表明iHog在Me、Mp和Mc中均有表达,以Me表达量最高,Mc表达量最低,差异显著(P0.05)。我们推测iHog的DNA序列的甲基化修饰抑制了该基因在Mc组织中的表达。综上研究结果表明,马氏珠母贝外套膜3个区域的甲基化修饰水平不一,且DNA甲基化在基因表达调控中具有作用,这对深入研究珍珠贝生物矿化和免疫反应机制具有一定的参考意义。  相似文献   

18.
The objective of this study was to assess sperm DNA longevity in blue mussels (Mytilus edulis) using a dynamic assessment of sperm DNA fragmentation (SDF) after sperm activation. Mature blue mussels (n = 57) in Vigo (Galicia, Spain) were obtained, specifically rope farmed blue mussels (n = 38) and wild blue mussels (n = 19). After the sperm collection, a subsample was assessed for SDF (0 h), while the rest of the sample was incubated for 6, 9, 12, 24 and 48 h at 15°C, assessing each time point using the Sperm‐Halomax kit (Halotech DNA, Madrid, Spain). The Kaplan–Meier estimator, log‐rank (Mantel–Cox) test and Mann–Whitney U‐test were used for statistical analyses (spss v. 16.0), α = 0.05. The rate of SDF (r‐SDF) between rope farmed and wild blue mussels over 0–6 h incubation was not significantly different (P = 0.278), but was for 6–24 h (P = 0.004). Differences in r‐SDF were observed when comparing the means between the two groups (P < 0.0001). Individual differences in r‐SDF existed among the rope farmed (P < 0.0001) and wild blue mussels (P < 0.0001). Wild blue mussels presented a higher DNA longevity than the farmed blue mussels. Selection of blue mussel males with a low level of sperm DNA damage and greater sperm DNA longevity may result in better fertilization and seed production.  相似文献   

19.
用纯化栉孔扇贝(Chlamys farrei)急性病毒性坏死症病毒(AVNV)免疫兔子,以兔抗血清为一抗,荧光标记的羊抗兔抗体为二抗,采用冰冻切片技术,建立了栉孔扇贝AVNV的间接免疫荧光检测方法。用该方法分析栉孔扇贝和海湾扇贝(Argopecten irradians)体内AVNV感染强度,并对感染率进行统计。结果表明,栉孔扇贝的肾脏、肝胰腺中,AVNV阳性信号最强,呈现中度到重度感染,其AVNV感染率100%。鳃丝、性腺及闭壳肌中未检测到阳性信号。在海湾扇贝的肝胰腺及肾中AVNV的感染率也较高。提示AVNV感染扇贝的靶器官主要是肾脏、肝胰腺。对与栉孔扇贝同一海区养殖的海湾扇贝在栉孔扇贝发病期间存活率较高的原因进行了讨论。  相似文献   

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