Complete nucleotide sequence of a cDNA encoding a myosin heavy chain from mantle tissue of scallop Patinopecten yessoensis

ABSTRACT: It has been reported that the amino acid sequences of striated and catch muscle myosin heavy chains from two scallop species ( Argopecten irradians and Placopecten magellanicus ) are almost identical, but that the ATPase activities between these myosins vary several-fold. These myosin sequences have been useful for identifying the region that modulates the ATPase activity of scallop myosin. In the present study, a cDNA encoding a myosin heavy chain was isolated from the mantle tissue of scallop Patinopecten yessoensis . The cDNA is composed of 6067 base pairs (bp) including an open-reading frame of 5841 p, which encodes an amino acid sequence of 1947 residues. The deduced amino acid sequence of P. yessoensis mantle myosin had a high identity of 90%, 92%, and 91% to P. magellanicus , A. irradians , and Pecten maximus striated muscle myosins, respectively. Interestingly, while the deduced amino acid sequences of around adenosine triphosphate-binding and actin-binding sites of the mantle myosin are homologous to those of A. irradians striated muscle myosin, the subfragment 2 hinge region and the non-helical tail region are similar to those of catch muscle myosin.     

cDNA clonings of shell matrix proteins from scallop shell

ABSTRACT:   A cDNA encoding the matrix shell protein from scallop (MSP-2) has been cloned. Comparison of the sequence of MSP-2 with the GenBank database revealed the sequence displays a high degree of homology with MSP-1, which was recently identified in the shell of the scallop Patinopecten yessoensis . Matrix shell protein-2 contains only 323 amino acids, while MSP-1 comprises 824 amino acids. The structural difference between MSP-1 and MSP-2 is mainly the presence or absence of the tandem repeat arrangement. In addition, multiple isoforms of MSP-2 were found. These isoforms share a high degree of sequence similarity to each other, suggesting that the MSP-2, MSP-2 isoforms and MSP-1 may constitute the same family. The MSP-2 isoforms in scallop shell may be responsible for the formation of molluscan shells.     

养殖密度对虾夷扇贝在浙江南麂海区生长的影响

通过设计D1、D2和D3 3个密度组,每组设3个平行笼,养殖密度分别为5、10和15ind·层^-1,研究养殖密度对虾夷扇贝（Patinopecten yessoensis）生长的影响。结果显示,随养殖密度的提高,壳高（SH）、体质量（BM）、相对壳高生长率（GRH）、相对体质量生长率（GRM）、日增壳高（DHG）、目增体质量（DMG）、体质量与壳高比及特定生长率（SGRSH和SGRBM）等参数不断下降,养殖密度对其影响显著（P〈0．05）。体质昔与壳高随生长时间分别呈现指数生长曲线与逻辑斯谛生长曲线。养殖密度对6月份的相对状态指数（RCF）及5月底的死亡率存存湿著影响（P〈0．05）,分析结果表明,10ind·层^-1为适宜的养殖密度。该研究为今后南方海Ⅸ虾夷扇贝规模化商业养殖提供参考.     

捕后处置对活品底播虾夷扇贝生化代谢的影响

为了研究干藏与湿藏对活品虾夷扇贝生化代谢的影响,探讨捕后干露、碰撞和温度3个胁迫因素对活品虾夷扇贝生化代谢的影响。将采捕后的虾夷扇贝分别进行干藏和湿藏处置,干藏采取冰藏方式,保藏时间为4 d;湿藏采取循环海水方式,同时施加温度与碰撞2种胁迫因素作对比,保藏时间为7 d。以闭壳肌中糖原、ATP及其关联物、AEC值、p H值和水溶性蛋白为指标,对4个处置组的活品虾夷扇贝进行跟踪分析。干藏条件下扇贝p H值、ATP含量和AEC值均迅速下降,生理状态快速恶化,短期内(1~3 d)扇贝陆续死亡。湿藏(5和10°C)条件下,扇贝闭壳肌可保持较高的ATP含量、AEC值和p H值,且5°C较10°C湿藏条件下糖原含量更高;相同温度下(10°C),增加碰撞会导致扇贝ATP含量和AEC值下降,糖原消耗增多。研究表明,捕后处置方式与活品虾夷扇贝的生化代谢有密切联系,干露是虾夷扇贝活品流通过程中比较突出的胁迫因素,温度和碰撞也会在一定程度上影响扇贝的生理状态。     

虾夷扇贝家系的建立及不同家系的早期生长研究

采用♂：♀=1：3的交配方法建立了33个虾夷扇贝Patinopecten yessoensis全同胞家系,对其壳长、壳高、壳宽和活体重进行了周年观测,对家系间和家系内的生长变异进行了比较分析。结果表明,不同家系间在不同性状上差异极显著。在同批家系中,06S-1、06S-24和06S-25号等家系壳高的生长较快,优势明显;06S-1、06S-25、06S-18和06S-30号等家系活体重的生长速度较快,差异显著;33个家系壳高月增长量大小顺序与活体重月增长量变化规律相近;06S-24、06S-25、06S-30、06S-1和06S-18号家系在各时期各性状上均表现出了明显的生长优势,是生长性状优良的家系,而06S-2和06S-5号家系生长一直最慢,各性状的表现也最差。全同胞家系间生长的差异性,表明家系间差异较显著,遗传变异资源丰富,可为后期的选育提供大量选种材料。     

温度对虾夷扇贝普通养殖群体和选育新品种海大金贝呼吸代谢的影响

2012年5月和9月,2013年3月和6月,在自然水温条件下,采用呼吸瓶法比较了不同温度(5.6℃、10.5℃、14.4℃、21.2℃)下普通养殖虾夷扇贝(Patinopecten yessoensis)和虾夷扇贝选育新品种海大金贝(Haida golden scallop)耗氧率和排氨率的影响。结果表明,在实验设置水温范围内(5.6~21.2℃),普通虾夷扇贝和海大金贝的耗氧率表现出相似的变化趋势。在温度达到14.4℃之前,实验贝耗氧率随温度的升高而增大,而在14.4℃后,则随温度的升高而减小。两种贝最大耗氧率分别为1.67 mg/(g·h)和1.27 mg/(g·h),其中在5.6℃和14.4℃海大金贝耗氧率显著小于普通虾夷扇贝(P0.05);10.5℃和21.2℃时,两组贝类的耗氧率差异不显著(P0.05)。普通虾夷扇贝和海大金贝排氨率随温度变化呈现出不同的趋势。前者从5.6℃开始,随温度的升高,排氨率缓慢升高,水温为14.4℃时达到最大值,为0.063 mg/(g·h),然后逐渐降低,14.4℃水温的排氨率显著大于10.5℃和21.2℃(P0.05);而从5.6℃到10.5℃,后者的排氨率逐渐降低,10.5℃时达到最低值,为0.029 mg/(g·h),然后随温度升高缓慢升高,到21.2℃达到最高值。海大金贝组在温度条件为5.6℃和21.2℃时排氨率高于普通虾夷扇贝组(P0.05);而水温为14.4℃时,普通虾夷扇贝组排氨率显著高于海大金贝组(P0.05);10.5℃时两者排氨率差异不显著(P0.05)。两实验组耗氧率Q_10系数均随温度的升高而降低。O/N结果表明,普通虾夷扇贝在本次研究的设定温度区间内以消耗脂肪和碳水化合物为主;海大金贝以消耗蛋白质为主,当温度逐渐升高,转化为以消耗脂肪和碳水化合物为主,当水温达到较高的水平,又转换为以消耗蛋白质为主。     

虾夷扇贝肌原纤维蛋白热稳定性的季节性差异

为探究虾夷扇贝原料特性的季节性差异,分别对春(4月)、夏(7月)、秋(10月)和冬(1月)产鲜活虾夷扇贝的商品属性及闭壳肌ATP和ATPase分布进行了分析比较;以肌原纤维蛋白(myofibrillar protein,Mf)Ca²⁺ATPase活性为检测指标,重点探索闭壳肌蛋白热稳定性的季节性差异,并对闭壳肌2种肌肉即横纹肌和平滑肌的蛋白热稳定性做了对比分析。结果显示,春季虾夷扇贝最肥满,生殖腺占总重的比率高达6.96%,其他3个季节仅为1.79%～2.95%。主要可食部位闭壳肌的蛋白质、脂肪和ATP的含量季节性差异不明显,闭壳肌富含蛋白质,含量占干基的75%～80%,脂肪含量很低,仅为0.63%～1.00%;值得关注的是,在春、夏2个季节闭壳肌总糖含量分别为11.14%和13.84%,显著高于秋、冬2季,而冬季含量最低,仅为2.67%。横纹肌ATP含量在各个季节都高于平滑肌,二者分别为4.3～5.0μmol/g和1.2～1.6μmol/g。横纹肌与平滑肌的ATP及其关联物含量的季节性差异均不明显。此外,横纹肌与平滑肌具有相似的热稳定性,在30和35℃下加热30...     

微酸性电解水对活品虾夷扇贝存活率的影响及杀菌效果

以活品虾夷扇贝(Patinopecten yessoensis)为实验对象,用不同理化性质的微酸性电解水(slightly acidic electrolyzed water,SAEW)处理受试扇贝,并用副溶血性弧菌人工浸染经微酸性电解水处理过的扇贝,检测其存活率、微酸性电解水杀菌效果、副溶血性弧菌的变化规律,以及虾夷扇贝在不同电解水处理阶段的菌相。结果显示,微酸性电解水处理1 min、2 min、4 min,扇贝存活率均为80%,处理8 min存活率为82%,均高于染菌组和对照组;电解水处理时间与细菌总数和大肠菌群呈显著负相关(P0.05),与处理时采用的电解水有效氯浓度没有显著性关系(P0.05)。采用电解水处理8 min后,活品虾夷扇贝体内染上的副溶血性弧菌数量从1 100 MPN/g降至28 MPN/g。扇贝初期主要菌群为假单胞菌和弧菌,在经过微酸性电解水处理后,其体内的菌群趋于复杂化,优势菌群有所改变,细菌总数有所下降。24 h后经电解水处理或者未处理的虾夷扇贝体内优势腐败菌均为假单胞菌。研究表明,SAEW在虾夷扇贝净化中具有一定的应用潜力。     

Culture of mantle epithelial cells expressing shell matrix proteins from scallop Patinopecten yessoensis

ABSTRACT:   In molluscs, mantle epithelial cells secrete organic matrix proteins to form shells. In this study, we established a culture of mantle epithelial cells by using the mantle pallial layer of scallops. We aimed to identify the mantle epithelial cells expressing scallop shell matrix proteins and establish a culture system of epithelial cells. After the mantle pallial layer was carefully isolated from the mantle tissue, explant culture was performed at 4°C. Most cells that migrated from the explant tissue were round cells. Most of the adhered cells retained round morphology, while some of the cells adhered to the dish and showed morphology similar to that of epithelial-like and fibroblast-like cells. When the cultured cells were immunostained with a polyclonal antibody against the shell matrix protein, the antibody recognized many of the adhered cells. An estimation of the number of epithelial cells revealed that approximately 70% of the adhered cells were epithelial cells. This is the first report to describe epithelial cells in cultured mantle cells, which express shell matrix proteins. This culture system may be a useful method for characterization of the mantle epithelial cells.     

Discovery of host defence genes in the Japanese scallop Mizuhopecten yessoensis Jay by expressed sequence tag analysis of kidney tissue

The Japanese scallop Mizuhopecten yessoensis is one of the most important aquaculture mollusca in Japan and China. In the present study, a high‐quality cDNA library of the Japanese scallop was constructed from the kidney tissue. A total of 2919 expressed sequence tags longer than 100 bp were generated from this library. A cluster of 1440 unique sequences, which consisted of 258 contigs and 1182 singletons, was revealed. Based on blast searches, 882 (61.3%) genes had significant (E‐value <1e–5) matches to known sequences in public databases. Among them, >70 genes were involved in stress response, immunity and apoptosis. These results expanded our knowledge of the genetics and physiology of the Japanese scallop, and provided a useful resource for gene discovery for further research of this species.     

cDNA cloning and expression analysis of the myosin heavy chain (MYH) gene of the mandarin fish Siniperca kneri

In this study, we applied RT-PCR and cDNA cloning techniques to clone myosin heavy chain (MYH) cDNA from muscle tissues of the mandarin fish Siniperca kneri . The cDNA was determined to be of 6987 base pairs in length, encoding a peptide of 1937 amino acids (Genbank accession no. EF446616). A search of encoded protein sequences in the NCBI conserved domain database indicated the presence of all known protein domains for MYH proteins, i.e. the myosin motor domain in the N-terminal region, the DIL domain at the C-terminus, and the ATPase domain. The MYH gene and its protein were expressed predominantly in muscle tissues and weakly in cardiac tissues. Developmentally, the MYH gene was first expressed in the muscle formation stage and continued later on. Our work provided a novel mypsin heavy chain gene sequence in fish biology and the results indicate that the MYH gene and the protein it encodes are important for the growth and development of the mandarin fish, as well as its muscle characterization.     

鲫血清转铁蛋白cDNA片段的克隆与序列分析

取体重约300g鲜活鲫(Carassius auratus)的肝组织进行总RNA提取。从GenBank数据库查询已发表的8种鱼转铁蛋白cDNA或基因序列,根据铁离子结合转运功能位点,设计并合成了2对引物P1、P4以及P2、P3,克隆出鲫血清转铁蛋白cDNA中的从大约550bp至1450bp的核心片段,长度为0．9kb。同时,比较了几种鱼血清转铁蛋白cDNA序列的同源性,并推导出鲫血清转铁蛋白cDNA核心片段的氨基酸序列,并分析了该氨基酸序列的结构和特性。     

鲫血清转铁蛋白cDNA克隆及系统发育进化序列分析

鱼类血清转铁蛋白是鱼类血清中一种非血红素结合铁的β—球蛋白。从GenBank数据库查询发表的鱼类转铁蛋白cDNA或基因序列,根据铁离子结合及转运功能位点,设计并合成了两对引物P1、P4以及P2、P3,克隆出鲫血清转铁蛋白cDNA中的核心片段,长度为866bp。再根据克隆出的核心片段分别设计上游及下游两对引物P5、P6以及P7、P8,随后用RACE方法分别克隆出鲫血清转铁蛋白cDNA的5’端(787bp)和3’端(1081bp)以及全长cDNA,最后在计算机上排列出鲫血清转铁蛋白全长cDNA,长度为2444bp。比较了14种鱼血清转铁蛋白cDNA序列的同源性,其同源性在30％～80％之间,结果显示鲤科鱼类(鲫、银鲫、鲤及斑马鱼等)具有很近的亲缘关系;同时进行了系统发育进化分析,证实了鱼类血清转铁蛋白进化的保守性和氨基酸序列的高度同源性。     

厚颌鲂(Megalobrama pellegrini)神经肽Y cDNA克隆和序列分析

利用RT-PCR的方法获得了厚颌鲂(Megalobrama pellegrini)NPY基因的编码序列。结果显示:厚颌鲂NPY基因长度为302 bp,包含了NPY基因291 bp的整个开放阅读框,共编码96个氨基酸,其分子量预测值为10987.4,理论等电点为5.72。通过ClustalX软件,将厚颌鲂与21个物种NPY的氨基酸序列进行比对,再结合Blastp分析的结果发现,厚颌鲂NPY氨基酸序列与鲤、中华倒刺鲃、鲫、斑马鱼、岩原鲤等鲤科鱼类的同源性最高,其同源性分别为100%、97%、95%、93%、93%。根据包括厚颌鲂在内的22个物种的NPY氨基酸序列的同源性构建进化树,其同源性与各鱼种分类地位一致。     

条斑紫菜细胞质甘油醛-3-磷酸脱氢酶的cDNA克隆和序列分析

利用已知甘油醛-3-磷酸脱氢酶(GAPDH)的特征性保守区域获得的探针筛选条斑紫菜cDNA文库,获得了1个全长的cDNA克隆GAP2,编码细胞质甘油醛-3-磷酸脱氢酶(GAPC)。其中GAP2 cDNA插入片段为1596bp,包括一个1008bp的开放阅读框编码336个氨基酸的细胞质甘油醛-3-磷酸脱氢酶(GAPC)蛋白。将获得的GAPC蛋白的氨基酸序列与其它物种的GAPDH序列进行多重序列比较后,发现条斑紫菜的GAPC氨基酸序列包含了4个高度保守的结构域。条斑紫菜GAPC的cDNA序列中GC含量很高,为64．2％,与紫菜EST分析中得到的结果(65．2％)近似。通过GAPDH的分子发育进化分析,对红藻的分类地位进行了初步探讨,条斑紫菜的细胞质甘油醛-3-磷酸脱氢酶的cDNA序列已登记到GenBank数据库,序列号分别为：AY273820。