排序方式: 共有43条查询结果,搜索用时 15 毫秒
11.
利用基因枪法将Tagsk1基因导入敏盐小麦成熟胚愈伤组织提高其耐盐性的研究 总被引:2,自引:1,他引:1
以Tagsk1(TriticumasetiumL.glycogen synthase kinase1)基因的cDNA的碱基序列为基础,设计特异引物由小麦耐盐突变体RH8706-49基因组DNA进行扩增后,得到来自于基因组的Tagsk1基因。采用基因枪法,利用携带该基因的双元表达载体pBI121-gsk1转化敏盐小麦H8706-34和中国春的成熟胚愈伤组织,经Kanamycin和0.5%NaCl筛选获得耐盐愈伤组织。这些被转化的愈伤组织表现出较高的耐盐性,并且能够在含盐培养基上进一步分化出根和芽。 相似文献
12.
Patrick Constantinescu Kati Kovacevic Giel J.C.G.M. Bosman Ronald Sluyter 《生物化学与生物物理学报:生物膜》2010,1798(9):1797-1804
Extracellular ATP induces cation fluxes in and impairs the growth of murine erythroleukemia (MEL) cells in a manner characteristic of the purinergic P2X7 receptor, however the presence of P2X7 in these cells is unknown. This study investigated whether MEL cells express functional P2X7. RT-PCR, immunoblotting and immunofluorescence staining demonstrated the presence of P2X7 in MEL cells. Cytofluorometric measurements demonstrated that ATP induced ethidium+ uptake into MEL cells in a concentration-dependent fashion and with an EC50 of ∼ 154 μM. The most potent P2X7 agonist 2′- and 3′-0(4-benzoylbenzoyl) ATP, but not ADP or UTP, induced ethidium+ uptake. ATP-induced ethidium+ and YO-PRO-12+ uptake were impaired by the P2X7 antagonist, A-438079. A colourmetric assay demonstrated that ATP impaired MEL cell growth. A cytofluorometric assay showed that ATP induced MEL cell death and that this process was impaired by A-438079. Finally, cytofluorometric measurements of Annexin-V binding and bio-maleimide staining demonstrated that ATP could induce rapid phosphatidylserine exposure and microparticle release in MEL cells respectively, both of which were impaired by A-438079. These results demonstrate that MEL cells express functional P2X7, and indicate that activation of this receptor may be important in the death and release of microparticles from red blood cells in vivo. 相似文献
13.
Grassi G Coceani N Farra R Dapas B Racchi G Fiotti N Pascotto A Rehimers B Guarnieri G Grassi M 《International journal of nanomedicine》2006,1(4):523-533
We studied the mechanism governing the delivery of nucleic acid-based drugs (NABD) from microparticles and nanoparticles in zero shear conditions, a situation occurring in applications such as in situ delivery to organ parenchyma. The delivery of a NABD molecule from poly(DL-lactide-co-glycolide) (PLGA) microparticles and stearic acid (SA) nanoparticles was studied using an experimental apparatus comprising a donor chamber separated from the receiver chamber by a synthetic membrane. A possible toxic effect on cell biology, as evaluated by studying cell proliferation, was also conducted forjust PLGA microparticles. A mathematical model based on the hypothesis that NABD release from particles is due to particle erosion was used to interpret experimental release data. Despite zero shear conditions imposed in the donor chamber, particle erosion was the leading mechanism for NABD release from both PLGA microparticles and SA nanoparticles. PLGA microparticle erosion speed is one order of magnitude higher than that of competing SA nanoparticles. Finally, no deleterious effects of PLGA microparticles on cell proliferation were detected. Thus, the data here reported can help optimize the delivery systems aimed at release of NABD from micro- and nanoparticles. 相似文献
14.
Chang-Moon?Lee Seung?Lim Gwang-Yun?Kim Doman?Kim Dong-Woon?Kim Hyun-Chul?Lee Ki-Young?LeeEmail author 《Biotechnology and Bioprocess Engineering》2004,9(6):476-481
The aim of this study was to formulate a sustained release system for indomethacin (IND) with rosin gum obtained from a pine
tree. Rosin microparticles were prepared by a dispersion and dialysis method without the addition of surfactant. In order
to investigate the influence of solvents on the formation of colloidal microparitcles, various solvents like ethanol, DMF,
DMAc, and acetone were used. The rosin microparticles containing IND were characterized by X-ray diffractometry (XRD) and
differential scanning calorimetry (DSC). The morphologies of rosin microparticles observed by scanning electron microscopy
(SEM) were spherical. The solvents used to dissolve rosin significantly affected the drug content and drug release rate of
IND. The release behaviors of IND from the rosin microparticles were dependent on the drug content and size of the particles.
Rosin microparticles with a higher drug content and of a larger particle size had a slower drug release rate. Also, the IND
release rate from the rosin microparticles could be regulated by the rosin content in the microparticles. From these results,
rosin microparticles have the potential of being used as a sustained release system of IND. 相似文献
15.
Oxyphor R2 and G2: phosphors for measuring oxygen by oxygen-dependent quenching of phosphorescence 总被引:1,自引:0,他引:1
Microparticles in the circulation activate the coagulation system and may activate the complement system via C-reactive protein upon conversion of membrane phospholipids by phospholipases. We developed a sensitive and reproducible method to determine the phospholipid composition of microparticles. Samples were applied to horizontal, one-dimensional high-performance thin-layer chromatography (HPTLC). Phospholipids were separated on HPTLC by chloroform:ethyl acetate:acetone:isopropanol:ethanol:methanol:water:acetic acid (30:6:6:6:16:28:6:2); visualized by charring with 7.5% Cu-acetate (w/v), 2.5% CuSO(4) (w/v), and 8% H(3)PO(4) (v/v) in water; and quantified by photodensitometric scanning. Erythrocyte membranes were used to validate the HPTLC system. Microparticles were isolated from plasma of healthy individuals (n = 10). On HPTLC, mixtures of (purified) phospholipids, i.e., lysophosphatidylcholine, phosphatidylcholine (PC), sphingomyelin (SM), lysophosphatidylserine, phosphatidylserine, lysophosphatidylethanolamine, phosphatidylethanolamine (PE), and phosphatidylinositol, could be separated and quantified. All phospholipids were detectable in erythrocyte ghosts, and their quantities fell within ranges reported earlier. Quantitation of phospholipids, including extraction, was highly reproducible (CV < 10%). Microparticles contained PC (59%), SM (20.6%), and PE (9.4%), with relatively minor (<5%) quantities of other phospholipids. HPTLC can be used to study the phospholipid composition of cell-derived microparticles and may also be a useful technique for the analysis of other samples that are available only in minor quantities. 相似文献
16.
Tamika K. Samuel Jason W. Sinclair Katherine L. Pinter Iqbal Hamza 《Journal of visualized experiments : JoVE》2014,(90)
In this protocol, we present the required materials, and the procedure for making modified C. elegans Habituation and Reproduction media (mCeHR). Additionally, the steps for exposing and acclimatizing C. elegans grown on E. coli to axenic liquid media are described. Finally, downstream experiments that utilize axenic C. elegans illustrate the benefits of this procedure. The ability to analyze and determine C. elegans nutrient requirement was illustrated by growing N2 wild type worms in axenic liquid media with varying heme concentrations. This procedure can be replicated with other nutrients to determine the optimal concentration for worm growth and development or, to determine the toxicological effects of drug treatments. The effects of varied heme concentrations on the growth of wild type worms were determined through qualitative microscopic observation and by quantitating the number of worms that grew in each heme concentration. In addition, the effect of varied nutrient concentrations can be assayed by utilizing worms that express fluorescent sensors that respond to changes in the nutrient of interest. Furthermore, a large number of worms were easily produced for the generation of transgenic C. elegans using microparticle bombardment. 相似文献
17.
Transformation of Nonselectable Reporter Genes in Marine Diatoms 总被引:6,自引:0,他引:6
Angela Falciatore Raffaella Casotti Catherine Leblanc Chiara Abrescia Chris Bowler 《Marine biotechnology (New York, N.Y.)》1999,1(3):239-251
We report the genetic transformation of two marine diatoms by microparticle bombardment. The pennate diatom Phaeodactylum tricornutum was transformed with the bacterial gene Sh ble from Streptoalloteichus hindustanus, which confers resistance to the antibiotics phleomycin and zeocin. Transformants contained between 1 and 10 copies of the
exogenous DNA integrated into the genome by illegitimate recombination at apparently random locations. Transformation efficiencies
were around 10−6, and individual cell lines could be maintained at −80°C following cryopreservation. Also, P. tricornutum could be transformed simultaneously with two different plasmids, one containing the Sh ble gene and another containing the firefly luciferase gene (LUC) under control of a promoter derived from a fucoxanthin, chlorophyll a/c-binding protein gene (FCP). In these cotransformants, LUC activity was light inducible. The transient transformation of the centric diatom Thalassiosira weissflogii with the bacterial β-glucuronidase (GUS) gene has also been achieved using similar transformation technology. The availability of gene transfer protocols for marine
diatoms, together with a range of functional reporter genes and regulated expression systems, will permit molecular dissection
of their biology and allow an assessment of the biotechnological potential of these organisms.
Received April 13, 1998; accepted November 16, 1998. 相似文献
18.
In HIV infections, homoeostasis of T cells is dysregulated such that there is a depletion of CD4+ T cells and a progressive loss of naïve CD4+ and CD8+ T cells. Methodologies that can improve the function of some or all of these cells will likely enhance immune responsiveness in HIV infection. Interleukin‐7 (IL‐7) is a cytokine that has been shown to be critical in homeostatic expansion of naïve CD8+ and CD4+ cells in lymphopenic hosts, as well as regulating effector T cell to memory T‐cell transition and memory T‐cell homeostasis. In animal studies and clinical trials, repeated injections of IL‐7 are used to boost both CD4+ and CD8+ cell counts. Daily injections, however, are painful, inconvenient, and provide a frequent route for pathogen entry. We developed a poly (D ,L ‐lactide‐co‐glycolide; PLGA) microparticle controlled release system to administer IL‐7 in which a single injection of microparticles can provide therapeutic delivery of IL‐7. IL‐7 encapsulated PLGA microparticles were first synthesized using a water/organic/water double emulsion method, release from the particles was then optimized using in vitro release studies and therapeutic effectiveness was finally studied in animal studies. These PLGA microparticles showed effective delivery of IL‐7 for 1 week in vitro. These results were translated to in vivo delivery as well, which was followed for 9 days. Controlled release of IL‐7 in mice demonstrated biological activity in both CD4+ and CD8+ T cells in mice, which was consistent with previously reported results using daily injections. Biotechnol. Bioeng. 2012; 109:1835–1843. © 2012 Wiley Periodicals, Inc. 相似文献
19.
Statin decreases endothelial microparticle release from human coronary artery endothelial cells: implication for the Rho-kinase pathway 总被引:8,自引:0,他引:8
Tramontano AF O'Leary J Black AD Muniyappa R Cutaia MV El-Sherif N 《Biochemical and biophysical research communications》2004,320(1):34-38
OBJECTIVE: Elevated plasma levels of endothelial microparticles (EMPs) are associated with the presence of clinical atherosclerosis. Considering the anti-inflammatory properties of HMG-CoA reductase inhibitors on the endothelium, we studied the effect of fluvastatin on the release of EMPs in cultured human coronary artery endothelial cells (HCAEC). METHODS AND RESULTS: EMPs were generated in TNF-alpha-activated HCAECs. The absolute number of EMPs was enumerated using a novel two-color flow cytometric immunostaining technique with TruCount beads as an internal reference. EMPs are defined as EC membrane vesicles (1-2 microm in size) with a characteristic immunophenotype. The addition of fluvastatin to TNF-alpha-activated HCAECs significantly suppressed EMP release. Fluvastatin suppressed TNF-alpha-induced Rho activation.The Rho-kinase inhibitor, Y-27632, reproduced the effect of statin. CONCLUSION: EMP release from TNF-alpha-activated HCAECs is suppressed by fluvastatin. In addition, the Rho/Rho-kinase may play an important role in modulating EMP release. 相似文献
20.
Sibel Yel İsmail Dursun Feyza Çetin Funda Baştuğ Sebahat Tülpar Ruhan Düşünsel 《Biomarkers》2018,23(6):558-562
Objective: Endothelial microparticles (EMPs) are considered as markers of endothelial dysfunction. In this study, we aimed to examine whether there is endothelial dysfunction in children with familial Mediterranean fever (FMF), hypothesizing that endothelial dysfunction would be present especially with acute-phase response in the active period of the disease.Methods: This cross-sectional study included 65 FMF patients (41 attack free, 24 attack period) and 35 healthy controls. Circulating EMPs, serum amyloid A (SAA), and other inflammation markers were measured in all groups. Circulating EMPs were measured using flow cytometry. Study groups were compared for circulating EMP and inflammatory markers. The relationship between EMPs and the activation of the disease was evaluated.Results: The levels of CD144+ and CD146+ EMPs in the FMF attack period group were significantly higher than those of the control group (p?0.05). The levels of inflammation markers in the attack period group were significantly higher than those of the control and attack-free groups (p?0.05). In the FMF attack group, the CD144+ and CD146+ EMP were significantly correlated with CRP.Conclusions: Our results suggest that endothelial damage is present especially in the active period of the disease in children with FMF. The endothelial dysfunction becomes an overt parallel with inflammation. 相似文献