Cloning and characterization of 37 kD laminin receptor precursor in pearl oyster, Pinctada fucata

A 1063 bp cDNA clone encoding a putative 37 kDa laminin receptor precursor （37 kDa LRP） is isolated from the mantle tissue of pearl oyster, Pinctadafucata. The amino acid sequence predicted from the cDNA sequence is 301 residues long, with a calculated molecular mass of 33.5 kDa. RT-PCR analysis shows that 37 kDa LRP mRNA is especially highly expressed in the mantle while widely expressed in several tissues. In situ hybridization analysis reveals that 37 kDa LRP is expressed in the outer epithelial ceils of the mantle edge, suggesting its involvement in cell proliferation and secretion in P. fucata. The identification and characterization of 37 kDa LRP in the pearl oyster will help us to further understand the signal transduction in the processes of mantle epithelial cell proliferation and tissue formation.     

Cloning and characterization of 37 kDa laminin receptor precursor in pearl oyster, Pinctada fucata

A 1063 bp cDNA clone encoding a putative 37 kDa laminin receptor precursor (37 kDa LRP) is isolated from the mantle tissue of pearl oyster, Pinctadafucata. The amino acid sequence predicted from the cDNA sequence is 301 residues long, with a calculated molecular mass of 33.5 kDa. RT-PCR analysis shows that 37 kDa LRP mRNA is especially highly expressed in the mantle while widely expressed in several tissues. In situ hybridization analysis reveals that 37 kDa LRP is expressed in the outer epithelial cells of the mantle edge, suggesting its involvement in cell proliferation and secretion in P. Fucata. The identification and characterization of 37 kDa LRP in the pearl oyster will help us to further understand the signal transduction in the processes of mantle epithelial cell proliferation and tissue formation.     

Cloning, Characterization, and Expression Analysis of Calreticulin from Pearl Oyster Pinctada fucata

Calreticulin is a unique calcium-binding protein with multiple functions mostly located in the sarcoplasmic/endoplasmic reticulum. A large amount of calcium is absorbed from the medium and transported to mineralization sites during biomineralization in pearl oyster. This paper describes the cloning of the full-length cDNA of calreticulin from Pinctada fucata, namely PCRT. PCRT encodes a deduced 414-amino acid protein, which includes a predicted 17- amino acid signal peptide and an endoplasmic reticulum retrieval sequence HDEL. The protein shows 63%-76% sequence identity and shares some common characteristics with calreticulins from other species. Semi-quantitative RT-PCR indicates that PCRT is ubiquitously expressed in all tissues tested with the highest expression in the hemolymph and the mantle. In situ hybridization analysis of PCRT in the mantle showed strong signals in the inner fold, the inner side of middle fold, and the inner side of outer fold of the mantle epithelium, All these results suggest PCRT might be involved in Ca^2＋ transport and storage during oyster biomineralization.     

育珠蚌外套膜组织培养适宜条件的研究

对育珠蚌外套膜细胞在组织培养过程中分泌珠质的适宜条件进行了研究。经过几年的实验和比较,筛选出了适宜于河蚌外套膜细胞生长、分泌的培养基及一套完善的操作方法,能使育珠蚌外套膜的细胞在体外人工培养条件下较旺盛地生长、繁殖、分泌,并产生大量的珍珠质.     

马氏珠母贝外套膜组织培养

用本实验室已建立的贝类组织培养技术对马氏珠母贝外套膜组织成功地进行了体外培养,在外套膜组织中最先迁出的是颗粒细胞,紧随其后为透明细胞,在培养到２０ｈ时,圆形的上皮细胞开始迁出,上皮细胞很快在组织块圆形形成生长晕,继而铺满整个培养瓶底面,培养４ｄ以后,上皮细胞开始分泌颗粒状的物质,这时的上皮细胞从形态上发为Ａ型和Ｂ型两类,Ｂ型上皮细胞含有许多颗粒物质,而Ａ型上皮细胞不含或含少量颗粒物质,反映了其合成     

背角无齿蚌组织培养细胞分泌的珍珠质的药理作用研究

利用背角无齿蚌的外套膜,进行离体组织培养,所得培养液及组织水解液,作用于大鼠离体子宫、兔离体小肠,其药理作用与珍珠质有效成分之一的牛磺酸相同,结果证明,背角无齿蚌外套膜培养细胞能分泌珍珠质,具有天然珍珠的药用有效成分及相同的活性.     

珍珠贝肉提取液抗衰作用的动物实验

在实验室检测马氏珍珠贝肉提取液对老龄小白鼠分辩学习及记忆保持能力的影响 ,并于行为实验结束后测定海马突触体内游离钙水平变化。小白鼠 1 6～ 1 7月龄 ,体重 (65±4) g,雌雄兼用 ,共 30只 ,分为天然提取液组 1 0只 ,强化提取液组 (天然提取液 +牛磺酸 ) 1 0只 ,对照组 (饮用普通清水 ) 1 0只。平均每只小鼠每天服用量为 5 ml,共服用 40 d。珍珠贝肉天然提取液和牛磺酸强化提取液与对照组相比 ,都明显增强老龄鼠在 Y-迷宫中的分辩学习能力和学习速度 ,减少训练次数 (P <0 .0 5) ;对记忆保持率的提高具有促进作用 ,尤其是牛磺酸强化提取液使记忆保持率大幅度提高 (P <0 .0 1 )。天然提取液与强化提取液都一致降低鼠脑的海马突触体游离钙离子水平 ,提示这种提取液对衰老脑细胞的“钙超载”有拮抗效应     

Molecular Cloning and Distribution of a Plasma Membrane Calcium ATPase Homolog from the Pearl Oyster Pinctada fucata

Plasma membrane calcium ATPaso (PMCA) plays a critical role in transporting Ca2 out of the cy- tosol across the plasma membrane which is essential both in keeping intracellular Ca2+ homeostasis and in biomineralization.In this paper we cloned and localized a gene encoding PMCA from the pearl oyster Pinctada fucata.This PMCA shares similarity with other published PMCAs within the functional domains.Reverse transcdption-polymerase chain reaction analysis shows that it is expressed ubiquitously.Furthermore,in situ hybridization reveals that it is expressed in the inner epithelial calls of the outer fold and in the outer epithelial calls of the middle fold,as well as the edge near the shell,which suggests that PMCA may be involved in calcified layer formation.The identification and characterization of oyster PMCA can help to further under-stand the structural and functional properties of molluscan PMCA,as well as the mechanism of maintaining Ca2+ homeostasis and the mechanism of mineralization in pead oyster.     

pEGFP-C1-MCH真核表达载体的构建及其在HEK293 细胞系中的表达

目的:构建pEGFP-C1-MCH真核表达载体,并将其转染入HEK293细胞中,筛选阳性细胞克隆,为研究MCH基因在能量代谢中的功能及机制提供细胞模型.方法:提取脑组织总RNA,反转录为cDNA,参照Genbank中提供的序列设计引物扩增MCH基因全长.再将该基因全长cDNA克隆至质粒pEGFP-C1,经菌落PCR筛选及双酶切和DNA测序鉴定,成功构建了含有目的基因MCH的重组质粒pEGFP-C1-MCH.并利用脂质体2000介导其转染HEK293细胞,用荧光显微镜和RT-PCR检测EGFP和MCH在细胞中的表达.结果:克隆的pEGFP-C1-MCH质粒序列中的MCH与Gen Bank相符;细胞转染72 h后,转染成功的细胞在荧光显微镜下表达较强的绿色荧光,MCH基因稳定表达.结论:pEGFP-C1-MCH真核表达载体的构建及其在HEK293细胞中的稳定表达,为研究MCH基因在能量代谢中的功能及作用机制提供了实验模型.     

可溶性人干细胞因子的原核表达和纯化

用Trizol提取人骨髓总RNA,通过RT-PCR扩增人干细胞因子DNA,克隆到pMD18-T载体中并测序．将其定向连入原核表达载体pET32a( ),获得了重组表达载体pET32a( )／hSCF,其cDNA长度为504bp,测序结果与已知序列吻合．然后在大肠杆菌BL21中经IPTG诱导表达,获得37kD的融合蛋白,约占菌体总蛋白量的30％．经Ni^2 -NTA树脂亲和层析柱纯化获得纯度大约90％的重组蛋白．     

Cloning, Characterization, and Expression Analysis of Calreticulin from Pearl Oyster Pinctada fucata

Calreticulin is a unique calcium-binding protein with multiple functions mostly located in the sar-coplasmic/endoplasmic reticulum.A large amount of calcium is absorbed from the medium and transported to mineralization sites during biomineralization in pead oyster.This paper describes the cloning of the full-length cDNA of calreticulin from Pinctada fucata,namely PCRT.PCRT encodes a deduced 414-amino acid protein,which includes a predicted 17- amino acid signal peptide and an endoplasmic reticulum retrieval sequence HDEL.The protein shows 63%-76% sequence identity and shares some common characteristics with calreticulins from other species.Semi-quantitative RT-PCR indicates that PCRT is ubiquitously ex-pressed in all tissues tested with the highest expression in the hemolymph and the mantle.In situ hybridiza-tion analysis of PCRT in the mantle showed strong signals in the inner fold,the inner side of middle fold,and the inner side of outer fold of the mantle epithelium.All these results suggest PCRT might be involved in Ca2+ transport and storage during oyster biomineralization.     

Effects on the early development of the pearl sac by cytochrome c, yolk lecithin and polyvinyplyrro

Mantle pieces ofHyriopisis cumingii was treated by three kinds of solution, These mantle pieces were inserted together with paraffin nucleuses into the connective tissue of three groups ofHyriopisis cumingii. These operated animals were cultured in a pool for a month. Several pearl sacs were put out and immersed by Bouin's solution after every five days. Sections of each animals were made by histological method. Those without treatment were in a control group. Observations of these sections showed that cytochrome c and yolk lecithin have an accelerated roles on pearl sac development and pearl layer secretion. No effects on pearl sac development by polyvinyplyrrolidone (PVP). Zheng Junying:born in May, 1966, Lecturer     

合浦珠母贝幼苗的正常生长

调查珍珠养殖合浦珠母贝人工培育幼苗的管养和生长情况,认为合浦珠母贝的幼苗生长正常,不存在“贝种退化”的影响,成活率和生长率的提高,明显是由于幼苗营养方法的改良。     

Cloning and identification of a novel human glioma differentiation related geneGDR1

In order to identify the genes associated with glioblastoma differentiation, some ESTs, expressed differentially in the control cell and the differentiated human glioblastoma cell line BT-325 induced by the all-trans retinoid acid, have been isolated by the method of DDRT-PCR. Of the 46 ESTs sequenced, 19 are from new genes. A full-length 1 535-bp cDNA, termed geneGDR1, has been isolated from the human cDNA library using the probe designed according to one of the novel ESTs, HGBB098. The open reading frame ofGDR1 gene encodes a putative protein containing 334 amino acid residues. Blast against the current GenBank DNA and protein sequence database did not reveal significant homology with any known proteins. RT-PCR shows thatGDR1 mRNA level increased in the differentiated BT-325 cells after being treated with RA. The different expression patterns ofGDR1 mRNA in human tissues have been detected through the multiple tissue Northern blot hybridization.     

Molecular cloning, identification and characteristics of NYD-SP9: Gene coding protein kinase presumably involved in spermatogenesis

Using cDNA microarray hybridization from a human testicular cDNA library, one gene exhibiting ten-fold difference at expression level between adult and embryo human testes was cloned and named NYD-SP9, which was believed to be involved in spermatogenesis. Southern blot hybridization results showed that NYD-SP9 expressed highly in testis but low in ovary. Protein motif analysis of this cDNA sequence revealed a cluster of phosphorylation sites, indicating its potential involvement in signal pathways during spermatogenesis. Furthermore, one transmembrane helix was predicted in N-terminal region, indicating that putative NYD-SP6 may be served as a transmembrane protein. The proximity of these potential phosphorylation sites to each other indicates that there may be interaction among these sites to regulate spermatogenesis. These findings suggested that protein kinase NYD-SP9 might play a role in male germ cell differentiation.     

漂白粉对三角帆蚌外套膜粘液细胞及珍珠囊细胞的影响

研究了不同浓度的漂白粉（0.5mg/L,1.0mg/L,1.5mg/L)处理后对三角帆蚌的影响,结果表明各浓度漂白粉对外套膜和珍珠囊细胞的超微结构均有不同程序损害,表现为粗面内质网脱颗粒,网腔扩张,线粒体基质电子密度改变等,其中1．5mg/L漂白粉的影响最为严重,除发生上述变化外,外套膜内表皮粘液细胞中硫酸化粘液减少,酸性粘液增加,外表皮细胞中性粘液几乎消失,该浓度组的结缔组织中钙球体的数量也明显高于其它各组。