共查询到20条相似文献，搜索用时 31 毫秒
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Chen Chen Yang-yang Fan Xin Wang Fei Song Tao Jiang Ping Qian Shun-ming Tang Xing-jia Shen 《Journal of Zhejiang University. Science. B》2016,17(2):127-135
Based on bioinformatic analysis, we selected two novel microRNAs (miRNAs), bmo-miR-0001 and bmomiR-0015, from high-throughput sequencing of the Bombyx mori larval posterior silk gland (PSG). Firstly, we examined the expression of bmo-miR-0001 and bmo-miR-0015 in 12 different tissues of the 5th instar Day-3 larvae of the silkworm. The results showed that the expression levels of both bmo-miR-0001 and bmo-miR-0015 were obviously higher in the PSG than in other tissues, implying there is a spatio-temporal condition for bmo-miR-0001 and bmo-miR-0015 to regulate the expression of BmFib-L. To test this hypothesis, we constructed pri-bmo-miR-0001 expressing the plasmid pcDNA3.0 [ie1-egfp-pri-bmo-miR-0001-SV40] and pri-bmo-miR-0015 expressing the plasmid pcDNA3.0 [ie1-egfp-pribmo-miR-0015-SV40]. Finally, the BmN cells were harvested and luciferase activity was detected. The results showed that luciferase activity was reduced significantly (P<0.05) in BmN cells co-transfected by pcDNA3.0 [ie1-egfp-pri-bmomiR-0001-SV40] or pcDNA3.0 [ie1-egfp-pri-bmo-miR-0015-SV40] with pGL3.0 [A3-luc-Fib-L-3′UTR-SV40], suggesting that both bmo-miR-0001 and bmo-miR-0015 can down-regulate the expression of BmFib-L in vitro. 相似文献
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Transfection and expression of exogenous gene in laying hens oviduct in vitro and in vivo 总被引：2，自引：0，他引：2
Gao B Sun HC Song CY Wang ZY Chen Q Song HQ 《Journal of Zhejiang University. Science. B》2005,6(2):137-141
To examine whether or not the regulatory sequence of chicken ovalbumin gene can drive transgene expression specifically in hen oviduct, the authors constructed an oviduct-specific expression vector (pOV), containing 3.0 kilobases (kb) of the 5'-flanking sequence and 3.0 kb of the 3'-flanking sequence of the chicken 相似文献
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Hong-cui Liu Bing-qiang Yuan Shao-nan Li 《Journal of Zhejiang University. Science. B》2016,17(2):110-126
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MicroRNAs (miRNAs or miRs) are endogenous non-coding RNAs that negatively regulate gene expression by binding to the 3′ non-coding regions of target mRNAs, resulting in their cleavage or blocking their translation. miRNAs may have an impact on cell differentiation, proliferation, and survival, and their deregulation can be inclined to diseases and cancers, including thyroid tumors. The purpose of this review is to summarize the existing findings of deregulated miRNAs in different types of thyroid tumors and to exhibit their potential target genes, especially to demonstrate those involved in tumor invasion and metastasis. In addition, new findings of circulating miRNA expression profiles, single nucleotide polymorphism (SNP) in thyroid tumors, and the correlation of somatic mutations with deregulated miRNA expression in thyroid tumors were all included in this review. 相似文献
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Wen-biao CHEN Jian-rong HUANG Xiang-qi YU Xiao-cong LIN Yong DAI 《Journal of Zhejiang University. Science. B》2015,16(3):235-250,166
目的:寻找来源于Alport综合征患者与正常对照者的多能干细胞差异性表达的microR NA,并对差异性表达的microR NA靶基因进行预测。创新点:本研究不同于一般的试验标本,试验标本是来源于尿肾脏管细胞诱导而成的多能干细胞。基于Alport综合征是遗传疾病,我们对一遗传家系进行了系统的分析。寻找特异性的差异性表达microR NA及其靶基因,从基因水平对Alport综合征进行研究。方法:在前期工作中,成功地从实验者与对照者的尿肾脏管细胞诱导成多能干细胞。运用高通量测序技术分析并发现实验者与对照者之间差异性表达的microR NA。对差异性表达的microR NA靶基因进行预测,并进行靶基因聚集分析,研究靶基因主要参与的生物学过程。同时进行靶基因信号传导通路的分析,研究靶基因主要参与的细胞信号传导通路。结论:在实验组与对照组之间,发现了30个具有显著差异性表达的microR NAs,包括19个上调表达与11个下调表达。差异性表达的microR NA的靶基因主要聚集在细胞膜和细胞代谢过程;靶基因主要参与嘌呤代谢通路与丝裂原活化蛋白激酶(MAPK)通路。 相似文献
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Lang QL Zhou XC Zhang XL Drabek R Zuo ZX Ren YL Li TB Chen JS Gao XL 《Journal of Zhejiang University. Science. B》2011,12(2):116-125
A large number of plant microRNAs (miRNAs) are now documented in the miRBase, among which only 30 are for Solanum lycopersicum (tomato). Clearly, there is a far-reaching need to identify and profile the expression of miRNAs in this important crop under
various physiological and pathological conditions. In this study, we used an in situ synthesized custom microarray of plant
miRNAs to examine the expression and temporal presence of miRNAs in the leaves of tomato plants infected with Cucumber mosaic virus (CMV). Following computational sequence homology search and hairpin structure prediction, we identified three novel tomato
miRNA precursor genes. Our results also show that, in accordance with the phenotype of the developing leaves, the tomato miRNAs
are differentially expressed at different stages of plant development and that CMV infection can induce or suppress the expression
of miRNAs as well as up-regulate some star miRNAs (miRNA*s) which are normally present at much lower levels. The results indicate
that developmental anomalies elicited by virus infection may be caused by more complex biological processes. 相似文献
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Objective: To construct a eukaryotic expression plasmid pcDNA3.1 (-)-Humanin. Methods: The recombinant plasmid pGEMEX-1-Humanin was digested with restriction endonucleases BamH I and Hind III and the Humanin gene fragments, about 100 bp length, were obtained. Then the Humanin gene fragments were inserted into eukaryotic expression vector pcDNA3.1 (-) and the recombinant plasmids pcDNA3, l(-)-Humanin were identified by sequencing. Results: Recombinant plasmid DNA successfully produced a band which had the same size as that of the Humanin positive control. The sequence of recombinant plasmids accorded with the Humnain gene sequence. Conclusions: A eukaryotic expression plasmid of Humanin was successfully constructed. 相似文献
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吴建桥 《孝感职业技术学院学报》2007,10(3):105-109
miRNA是一类高度保守的非编码RNA,为21-25nt的单链型RNA(ssRNAs),由70-90nt的可形成发夹状的内源性转录前体pri-miRNA加工而成,miRNA与靶mRNA的3'-UTR碱基配对,通过抑制mRNA的翻译或直接使mRNA降解,在转录后水平使基因产生沉默,研究发现人类全部基因的三分之一都受到miRNA的调控,这表明miRNA分子实际上是基因调控网络中的核心成分。近来研究发现miRNA与人类的某些疾病密切相关。文章综述了miRNA的基因调控原理和临床研究进展。 相似文献
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Computational prediction of microRNA genes in silkworm genome 总被引：3，自引：0，他引：3
MicroRNAs （miRNAs） constitute a novel, extensive class of small RNAs （-21 nucleotides）, and play important gene-regulation roles during growth and development in various organisms. Here we conducted a homology search to identify homologs of previously validated miRNAs from silkworm genome. We identified 24 potential miRNA genes, and gave each of them a name according to the common criteria. Interestingly, we found that a great number of newly identified miRNAs were conserved in silkworm and Drosophila, and family alignment revealed that miRNA families might possess single nucleotide polymorphisms, miRNA gene clusters and possible functions of complement miRNA pairs are discussed. 相似文献
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To construct and identify further a recombinant of Adeno-associated virus and interferon-gamma for gene therapy, the full-length
IFN-γcDNA containing signal peptide was amplified by PCR, and then cloned into the pUC18. After screening, the fragment from
the positive clone was then subcloned into pwp19. After the correct recombinant was identified by digestion with SacI and
BamHI, it was transfected into lymphocyte cell line H9 mediated by calcium phosphate, and the expression of IFN-γ was detected
by RT-PCR and ELISA. The result showed that the IFN-γ were expressed in the H9 cells transfected with pwp/IFN-γ. The so constructed
recombinant plasmid pwp19/IFN-γ containing the full-length IFN-γ gene was expressed in mammalian cells.
Project (39570653) supported by NFSC. 相似文献
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目的：探讨增强型绿色荧光蛋白（enhanced green fluorescent protein, EGFP）在中华仓鼠卵巢细胞（CHO-DHFR）细胞中的作用。方法：将增强型绿色荧光蛋白基因的真核表达载体pcDNA3．1（＋）-EGFP，转染至培养的中华仓鼠卵巢细胞（Chinese Hamster Ovary, CHO-DHFR^- ）中。结果：成功表达并产生绿色荧光。结论：证明EGFP是一种良好的报告基因和筛选标志．为进一步研究应用最广泛的哺乳动物细胞表达系统一CHO细胞表达系统奠定了基础。 相似文献
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Xing-wei Xiang Rui Yang Lin Chen Xiao-long Hu Shao-fang Yu Cui-ping Cao Xiao-feng Wu 《Journal of Zhejiang University. Science. B》2012,13(2):111-117
In the late phase of Bombyx mori nucleopolyhedrovirus (BmNPV) infection, a large amount of polyhedra appear in the infected cell nucleolus, these polyhedra
being dense protein crystals protecting the incorporated virions from the harsh environment. To investigate whether the foreign
protein could be immobilized into the polyhedra of BmNPV, two recombinant baculoviruses were generated by a novel BmNPV polyhedrin-plus
(polh+) Bac-to-Bac system, designated as vBmBac(polh+)-enhanced green fluorescent protein (EGFP) and vBmBac(polh+)-LacZ, which can express the polyhedrin and foreign protein simultaneously. Light microscopy analysis showed that all viruses
produced polyhedra of normal appearance. Green fluorescence can be apparently detected on the surface of the vBmBac(polh+)-EGFP polyhedra, but not the BmNPV polyhedra. Fluorescence analysis and anti-desiccation testing confirmed that EGFP was
embedded in the polyhedra. As expected, the vBmBac(polh+)-LacZ polyhedra contained an amount of LacZ and had a higher β-galactosidase activity. Sodium dodecyl sulfate polyacrylamide
gel electrophoresis (SDS-PAGE) and Western blotting were also performed to verify if the foreign proteins were immobilized
into polyhedra. This study provides a new inspiration for efficient preservation of useful proteins and development of new
pesticides with toxic proteins. 相似文献
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Na Wang Cai-ying Jiang Ming-xing Jiang Chuan-xi Zhang Jia-an Cheng 《Journal of Zhejiang University. Science. B》2010,11(9):728-734
The piggyBac transposon has been long used to integrate foreign DNA into insect genomes. However, undesirable transgene expression can
result from random insertions into the genome. In this study, the efficiency of chimeric Gal4-piggyBac transposase in directing integration onto a DNA target plasmid was evaluated in cultured silkworm Bombyx mori Bm-12 and fruit fly Drosophila Schneider 2 (S2) cells. The Gal4-piggyBac transposase has a Gal4 DNA-binding domain (DBD), and the target plasmid has upstream activating sequences (UAS) to which
the Gal4 DBD can bind with high affinity. The results indicate that, in the Bm-12 and S2 cells, transpositional activity of
Gal4-piggyBac transposase was increased by 4.0 and 7.5 times, respectively, compared to controls, where Gal4-UAS interaction was absent.
Moreover, the Gal4-piggyBac transposase had the ability of directing piggyBac element integration to certain sites of the target plasmid, although the target-directing specificity was not as high as
expected. The chimeric piggyBac transposase has the potential for use in site-directed transgenesis and gene function research in B. mori. 相似文献
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Li-rong Shen Mei-hui Ding Li-wen Zhang Wei-guang Zhang Liang Liu Duo Li 《Journal of Zhejiang University. Science. B》2010,11(5):342-349
Bee venom phospholipase A2(BvPLA2) is a lipolytic enzyme that catalyzes the hydrolysis of the sn-2 acyl bond of glycerophospholipids to liberate free fatty acids and lysophospholipids.In this work,a new BvPLA2(AccPLA2) gene from the Chinese honeybee(Apis cerana cerana) venom glands was inserted into bacmid to construct a recombinant transfer vector.Tn-5B-4(Tn) cells were transfected with the recombinant bacmid DNA for expression.Sodium dodecylsulfate-polyacrylamide gel electrophoresis(SDS-PAGE) analysis revealed a double band with molecular weights of 16 and 18 kDa.Products of hexahistidine AccPLA2 fusion protein accumulated up to 5.32% of the total cellular proteins.The AccPLA2 fusion protein was cross reactive with the anti-AmPLA2(BvPLA2 of the European honeybee,Apis mellifera) polyclonal serum.The reaction resulted in a double glycosylation band,which agrees with the band generated by the native AmPLA2 in Western blot analysis.The PLA2 activity of the total extracted cellular protein in the hydrolyzing egg yolk is about 3.16 μmol/(min·mg).In summary,the recombinant AccPLA2 protein,a native BvPLA2-like structure with corresponding biological activities,can be glycosylated in Tn cells.These findings provided fundamental knowledge for potential genetic engineering to produce AccPLA2 in the pharmaceutical industry. 相似文献
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增殖细胞核抗原(proliferating cell nuclear antigen,PCNA),也称周期蛋白或DNA聚合酶的辅助蛋白,是真核细胞合成所必需的核蛋白,在DNA复制中起重要作用。前期实验发现日本七鳃鳗肝脏cDNA文库的表达序列标签(Expressed Sequence Tag,EST)中存在与高等脊椎动物pcna基因同源的序列。提取日本七鳃鳗(Lampetra japonica)肝脏组织RNA,通过RT-PCR方法扩增七鳃鳗pcna基因,对其进行生物信息学分析,并将Lj-pcna基因成功构建到pGFP-N2真核表达载体上,重组质粒PGFP-N2-Lj-pcna转染人Hela细胞,荧光显微镜下观察有荧光蛋白的表达。日本七鳃鳗pcna基因的真核表达载体成功构建和转染,为探讨七鳃鳗pcna基因功能研究及其它七鳃鳗相关研究提供条件。 相似文献
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采用PCR方法扩增对虾白斑病毒核糖核苷酸还原酶基因,插入到pGEX-4T-2表达载体上构建出带有目的基因的重组质粒pGEX-4T-2RR,然后将重组质粒转化大肠杆菌.转化菌经IPTG诱导后大量表达重组蛋白,通过降低诱导温度获得可溶性表达的重组蛋白,采用Glutathione Sepharose 4B亲和层析对重组蛋白进行纯化,获得了高纯度的目的蛋白. 相似文献