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GH-IGF-I轴是鱼体体内一个重要的内分泌生理轴,主要调控鱼体的生长发育。用RT-PCR方法从加州鲈脑垂体和肝脏组织中分别扩增出加州鲈GH和IGF-I cDNA,克隆到pMD19 T-Vector上进行序列测定和分析。结果表明:1)加州鲈GH cDNA开放阅读框长为615 bp,编码204个氨基酸,其中信号肽17个氨基酸,成熟肽187个氨基酸。成熟肽中有四个保守的半胱氨酸残基(分别位于69,177,193,202),可形成两对二硫键。加州鲈GH氨基酸序列与蓝太阳鱼、斜带石斑鱼、金头鲷、虹鳟、鲤鱼、斑马鱼相比较,同源性分别为100%、97%、94%、66%、56%、53%。2)加州鲈IGF-I cDNA开放阅读框长为561bp,编码包括信号肽和B、C、A、D、E五个区域的186个氨基酸,形成成熟肽时,信号肽和E区域被切除。成熟蛋白的氨基酸序列与GenBank中已知的河鲈、三角鲂、金头鲷、舌齿鲈、斑马鱼的相比较,结果发现四个区域的保守性有差异,A区和B区保守性较高,C区和D区保守性较差。加州鲈GH和IGF-I cDNA的获得为进一步研究鱼体GH-IGF-I轴对生长发育的调控机制奠定了基础。 相似文献
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将大口黑鲈(Micropterus salmoide)肌肉生长抑制素(Myostatin,MSTN)前肽(MSTN-Pro)的cDNA定向克隆到真核表达载体pcDNA3.1(-)/mycHisB中,双酶切检测和测序鉴定证实,插入pcDNA3.1(-)/mycHisB载体中的片段为目的基因的核苷酸序列,MSTN基因前肽cDNA为正向插入,且重组质粒无错配或插入移位等突变。采用肌肉注射法将重组表达质粒注入大口黑鲈背部肌肉组织,在注射后第2天经RT-PCR检测到MSTN前肽基因mRNA的表达,第6天经免疫组化学检测到MSTN前肽蛋白的表达,第8天蛋白表达强度增强,对照组始终未检测到MSTN前肽基因的表达。 相似文献
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大口黑鲈抗菌肽hepcidin cDNA序列和结构分析 总被引：2，自引：0，他引：2
以大口黑鲈为材料,提取肝脏总RNA,经RT-PCR扩增出hepcidin cDNA的开放阅读框(ORF)及3′端非编码区序列,应用5′RACE方法得到大口黑鲈hepcidin cDNA5′末端。将所获得的两个片段分别克隆到T载体后进行测序,并拼接成大口黑鲈hepcidin全长cDNA。序列分析表明:大口黑鲈hepcidin全长cDNA为564bp,含有一个258bp的ORF,编码86个氨基酸残基,由信号肽(24个残基)、前肽(42个残基)和成熟肽(20个残基)3部分组成hepcidin前体。在前肽部分具有前肽转化酶典型的RX(K/R)R基元,成熟肽部分含有8个保守的半胱氨酸残基,可形成四个链内二硫桥,使β-折叠结构保持稳定。大口黑鲈Hepcidin与其他鱼类的同源性在29.7%~90.5%间,尤其是信号肽区域,与鳜、尼罗罗非鱼、真鲷、花鲈、黑鯛、金眼狼鲈仅有2~3个氨基酸的差别。 相似文献
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Shuo Li Chunmei Li Xubo Wang Yanan Wang Zhipeng Liu Teng Zhai Quanqi Zhang 《中国海洋大学学报(英文版)》2010,9(1):59-64
A novel immune-related gene was expressed in Japanese flounder (Paralichthys olivaceus) injected with Vibrio anguillarum. The complete cDNA contained a 169 bp 5’UTR, a 336 bp open reading frame (ORF) encoding 111 amino acids and a 556bp 3’UTR.
Six exons and five introns were identified in the PoIR2 gene. Blastp similarity comparison showed its encoding protein had 50% similarity to Danio rerio neuromedin S (NMS), but further alignment indicated they did not have NMS C-terminal conservational signature domain. So
it was not defined as an NMS homologue. Protein structure analysis indicated it had a 26aa signal peptide and was a secretory
pathway protein. RT-PCR demonstrated that the expression of PoIR2 was quickly induced and drastically increased in liver, kidney, spleen, gills, intestine, heart, and skeletal muscle after
infected with V. anguillarum. These results indicated that the PoIR2 might play some important role in Japanese flounder immune response system. This gene was named PoIR2 (P.olivaceus immune-related gene 2, GenBank accession number: EU224372). The mature PoIR2 peptide was expressed in BL21(DE3) pLysS using pET-32a(+) vector and a great part of the recombinant mature peptide existed
as soluble type. 相似文献
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四年来，在研究了大口黑鲈生物学特性的基础上，吸收了国内外的养殖经验，结合实际确立了在水泥池和池塘中繁殖鱼苗、培育鱼种和池塘主养、配养食用鱼的养殖技术。 相似文献
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Galectins,a family of ?-galactoside-binding proteins,participate in both innate immunity and adaptive immunity.This study identified one novel galectin-related protein from half-smooth tongue sole Cynoglossus semilaevis,which was designated as Cs GRP.The full-length c DNA of its encoding gene was 785 bp in length with a 528 bp open reading frame encoding a putative protein of 176 amino acids.The deduced Cs GRP contained a putative 131-aa galactoside-binding lectin domain and 3 critical residues responsible for carbohydrate binding(R~(93),W~(109) and R~(114)).Genomic structural analysis revealed that Cs GRP consisted of five exons and four introns.Cs GRP showed 68% similarity with Poecilia latipinna GRP and 67% similarity with Stegastes partitus GRP.Cs GRP showed the highest expression level in liver,although its expression was detected in all tested tissues.When challenged with Vibrio harveyi,the expression of Cs GRP was significantly down-regulated in liver(P 0.05).In addition,we found that in spleen and kidney of C.semilaevis,the Cp G island of Cs GRP showed significantly higher(P 0.05) methylation level in disease-resistant family of C.semilaevis(DR-Cs) than in disease-susceptible ones(DS-Cs).Our results suggested that Cs GRP may play important roles in the immune response of C.semilaevis.Moreover,DNA methylation patterns provided valuable data for understanding the relationship between epigenetic regulation and immunity,which would assist the animal genetic research and improve the animal breeding in the future. 相似文献
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大口黑鲈选育群体遗传结构的微卫星分析 总被引：1，自引：0，他引：1
采用微卫星标记技术分析不同世代大口黑鲈选育群体的遗传结构。利用11对微卫星引物对大口黑鲈第2～4代选育群体及南水群体(对照)共120个个体进行PCR扩增,共得到28个等位基因。每个位点获得2～3个等位基因,平均等位基因为2.54。第2代(F_2)、第3代(F_3)、第4代(F_4)及南水群体(CG)的平均多态信息含量(PIC)值分别为:0.423、0.419、0.386、0.366。与F_2相比,F_4的遗传多样性减少8.76%,但与CG相比,F_4仍具有较高的遗传多样性,表明大口黑鲈选育群体的遗传基础逐步趋向纯化,且仍有较大选育潜力。此外,尽管F_2与F_4的遗传距离(0.022)比F_2与F_3的(0.011)大,但世代间的F_(st)值(0.006～0.009)均小于0.05,且世代群体总遗传分化指数为0.013,表明选育群体已出现遗传分化,但分化程度较低。 相似文献
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双组分调控系统（Two-component Regulatory System）在致病菌的生长及毒力调控中起重要作用.克隆溶藻弧菌（Vibrio alginolyticus）HY9901 株组氨酸激酶PhoR 和反应调控因子PhoB 的全长基因,并对其进行生物信息学分析.序列分析结果显示,phoR（GenBank 登录号：KJ958404）全长1 299 bp,共编码432 个氨基酸;phoB（GenBank 登录号：KJ863646）全长690 bp,编码229 个氨基酸.构建PhoR/PhoB 的系统进化树,结果显示,溶藻弧菌PhoR/PhoB 与副溶血弧菌（Vibrio parahaemolyticus）、哈氏弧菌（Vibrio harveyi）有较近亲缘关系.利用SWISS-MODEL 软件,对PhoR/PhoB 中两个相对保守的功能域HATPase_c 和REC 进行同源建模,发现HATPase_c具有1 个ATP 结合位点,REC 具有4 个天冬氨酸活性位点. 相似文献
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用RT-PCR和RACE方法,从荷那龙罗非鱼(Oreochromis hornorum)垂体中克隆到生长激素促分泌素受体(GHSR)cDNA全序列。荷那龙罗非鱼GHSR基因具有GHSR-1a与GHSR-1b两个高度保守cDNA序列。GHSR-1a序列全长1 646 bp,包括225 bp的5′非编码区,266 bp的3′非编码区和1 155 bp的开放阅读框,编码384个氨基酸残基,具有7个跨膜结构域结构(transmembrane domains,TM);GHSR-1b序列全长1 877 bp,包括225 bp的5′非编码区,755 bp的3′非编码区和897 bp的开放阅读框,编码298个氨基酸残基,只具有前5个TM,在第6个TM的第4个氨基酸处开始缺失。将GHSR cDNA序列与基因组序列比较发现,这两种cDNA转录本来自同一个基因的不同变体。 相似文献
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According to the known sequence of iron stress-induced gene (isiAB operon), we cloned its 1.5 kb fragment by PCR, and used this fragment as integration homologous fragment. After several steps of subcloning donor DNA into theisiAB fragment, a donor plasmid pZL which could be integrated into the chromosomal DNA ofSynechococcus sp. PCC7942 was constructed. In order to express the heterologous gene at a high level through the integration platform system, we constructed the donor DNA by the following steps. We cloned the strong promoter (240 bp) of heat shock genegroESL operon fromSynechococcus sp. PCC7942 by PCR. Then subcloned the multiple cloning sites (MCS),rbcS polyA into the downstream of thegroESL promoter. The kanamycin resistance gene, as the marker gene, was also subcloned into the donor DNA. Thus, in the donor plasmid pZL, the integration homologous fragment and several expression elements, such asgroESL promoter, MCS,rbcS polyA terminator and kanamycin resistance gene, were all included.
After naturally transformed and introduced the donor plasmid pZL intoSynechococcus sp. PCC7942, as in the pZL, the donor DNA sequence is flanked by two DNA fragments (0.4 kb and 0.7 kb) homologous to theisiAB fragment ofSynechococcus sp. PCC7942, the homologous DNA can recombine with the chromosomal DNA. After screening by kanamycin, the transformants which integrated the heterologous DNA were selected. The efficiency of transformation is about 1×10−6. By southern blot analysis, it was confirmed that the donor DNA had been integrated into the chromosomal DNA ofSynechococcus sp. PCC7942, located on the site of theisiAB gene, and can be replicated with the chromosomal DNA.
相似文献17.
A muscle cDNA library of Chinese shrimp (Fenneropenaeus chinensis) was constructed with the SMARTTM cDNA Li- brary Construction Kit. The titer of optimal primary library was 7.7×105 pfu mL-1 and that of the amplified library was 3.0×109 pfu mL-1. The percentages of the recombinant clones of primary and amplified libraries were over 98%. The insert sizes were longer than 400 bp with an average of 1000 bp. A positive clone containing a 794 bp insert was sequenced and identified encoding fast skeletal troponin Ⅰ gene. This library provided a useful resource for the functional genomic research ofE chinensis. 相似文献
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Using shotgun sequencing data, the complete sequences of chloroplast 16S rRNA and tufA genes were acquired from native specimens of Bryopsis hypnoides (Qingdao, China). There are two group I introns in the 16S rRNA gene, which is structurally similar to that of Caulerpa sertularioides (Bryopsidales, Chlorophyta). The chloroplast-encoded tufA gene sequence is 1 230 bp long, very AT-rich (61.5%), and is similar to previously published 16S rRNA sequences of bryopsidinean algae. Phylogenetic analyses based on chloroplast 16S rRNA and tufA gene sequence data support previous hypotheses that the Bryopsidineae, Halimedineae, and Ostreobidineae are three distinct lineages. These results also confirmed the exclusion of Avrainvillea from the family Udoteaceae. Phylogenetic analyses inferred that the genus Bryopsis as sister to Derbesia; however, this clade lacked robust nodal support. Moreover, the phylogenetic tree inferred from rbcL GenBank sequences, combined with the geographical distributions of Bryopsis species, identified a strongly supportive clade for three differently distributed Asian Bryopsis species. The preliminary results suggesting that these organisms are of distinct regional endemism. 相似文献
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The myosin heavy chain (MyHC) is one of the major structural and contracting proteins of muscle. We have isolated the cDNA
clone encoding MyHC of the grass carp, Ctenopharyngodon idella. The sequence comprises 5 934 bp, including a 5 814 bp open reading frame encoding an amino acid sequence of 1 937 residues.
The deduced amino acid sequence showed 69% homology to rabbit fast skeletal MyHC and 73%–76% homology to the MyHCs from the
mandarin fish, walleye pollack, white croaker, chum salmon, and carp. The putative sequences of subfragment-1 and the light
meromyosin region showed 61.4%–80% homology to the corresponding regions of other fish MyHCs. The tissue-specific and developmental
stage-specific expressions of the MyHC gene were analyzed by quantitative real-time PCR. The MyHC gene showed the highest
expression in the muscles compared with the kidney, spleen and intestine. Developmentally, there was a gradual increase in
MyHC mRNA expression from the neural formation stage to the tail bud stage. The highest expression was detected in hatching
larva. Our work on the MyHC gene from the grass carp has provided useful information for fish molecular biology and fish genomics. 相似文献