Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “GFP”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 217 records · Page 12Linked to original sources
PubMed2001

Green fluorescent protein (GFP) transgenic fish and their applications.

The coupling of the GFP reporter system with the optical clarity of embryogenesis in model fish such as zebrafish and medaka is beginning to change the picture of transgenic fish study. Since the advent of first GFP transgenic fish in 1995, GFP transgenic fish technology have been quickly employed in many areas such as analyses of gene expression patterns and tissue/organ development, dissection of promoters/enhancers, cell lineage and axonal pathfinding, cellular localization of protein products, chimeric embryo and nuclear transplantation, cell sorting, etc. The GFP transgenic fish also have the potentials in analysis of upstream regulatory factors, mutagenesis screening and characterization, and promoter/enhancer trap. Our own studies indicate that GFP transgenic fish may become a new source of novel variety of ornamental fish. Efforts are also being made in our laboratory to turn GFP transgenic fish into biomonitoring organisms for surveillance of environmental pollution.

Animals↗
PubMed2002

Developmental and tissue-specific expression of CaMV 35S promoter in cotton as revealed by GFP.

The CaMV 35S promoter is the most commonly used promoter for driving transgene expression in plants. Though it is presumed to be a constitutive promoter, some reports suggest that it is not expressed in all cell types. In addition, the information available on its expression profile in all possible cell and tissue types and during early stages of development is incomplete. We present here a detailed expression profile of this promoter investigated using the green fluorescent protein (GFP) gene as a reporter system in cotton during embryo development, and in all the vegetative and floral cell and tissue types. GFP expression was not detected during the early stages of embryogenesis. The first perceptible GFP expression was observed in a small area at the junction of hypocotyl and cotyledons in embryos at around 13 days after anthesis. The GFP fluorescence progressively became stronger and expanded throughout the cotyledon and hypocotyl as embryo development advanced. After germination, varying levels of promoter activity were observed in all cell and tissue types in the hypocotyl, cotyledon, stem, leaf, petiole, and root. The promoter was also expressed in all floral parts. Although cotton pollen exhibited a low level of greenish autofluorescence, it was possible to discern GFP-dependent fluorescence in some of the pollen from all the T0 plants examined. Developing cotton fibers also exhibited GFP fluorescence suggesting that the 35S promoter was active in these specialized epidermal cells. Thus, we show that the expression of the 35S promoter was developmentally regulated during embryogenesis and that beyond a certain stage during embryogenesis, the promoter was expressed in most cell and tissue types in cotton albeit at different levels.

Caulimovirus↗
PubMed2003

Overexpression and mislocalization of a tail-anchored GFP redefines the identity of peroxisomal ER.

Peroxisomal ascorbate peroxidase (APX) sorts indirectly via a subdomain of the ER (peroxisomal ER) to the boundary membrane of peroxisomes in tobacco Bright Yellow 2 cells. This novel subdomain characteristically appears as fluorescent reticular/circular compartments distributed variously in the cytoplasm. Further characterizations are presented herein. A peptide possessing the membrane targeting information for peroxisomal APX was fused to GFP (GFP-APX). Transiently expressed GFP-APX sorted to peroxisomes and to reticular/circular compartments; in both cases, the GFP moiety faced the cytosol. Of particular interest, both homotypic and heterotypic aggregates of peroxisomes, mitochondria, and/or plastids were formed. The latter two organelles comprised the circular portion of the reticular/circular compartments, apparently as a consequence of oligomerization (zippering) of the GFP moieties after insertion into the outer membranes of the affected organelles. These results, coupled with the accumulation of endogenous peroxisomal APX in cytoplasmic, noncircular compartment(s) following treatment with brefeldin A, indicate that authentic peroxisomal ER is composed only of a reticular compartment(s). Equally important, the data show that overexpressed, membrane-targeted GFP fusion proteins have a propensity to form organelle aggregates that may lead to misinterpretations of sorting pathways of trafficked proteins.

Ascorbate Peroxidases↗
PubMed1996

Spatial dynamics of GFP-tagged proteins investigated by local fluorescence enhancement.

We describe a method of monitoring the spatial dynamics of proteins in intact cells by locally enhancing the blue excited fluorescence of green fluorescent protein (GFP) using a spatially focused ultraviolet-laser pulse. GFP fusion proteins were efficiently expressed by micro-electroporation of in vitro synthesized mRNA into adherent mammalian cells. We found that the diffusion coefficient of cycle 3 mutant GFP was 43 microns2/sec, compared to 4 microns2/sec for wild-type GFP, suggesting that cycle 3 GFP diffuses freely in mammalian cells and is ideally suited as a fusion tag. The local fluorescence enhancement method was used to study the membrane dissociation rate of GFP-tagged K-ras, a small GTP binding protein that localizes to plasma membranes by a farnesyl lipid group and a polybasic region. Our data suggest that K-ras exists in a dynamic equilibrium and rapidly switches between a plasma membrane bound form and a cytosolic form with a plasma membrane dissociation time constant of 1.5 sec.

Animals↗
PubMed2004

Soluble expression and characterization of a GFP-fused pea actin isoform (PEAc1).

A pea actin isoform PEAc1 with green fluorescent protein (GFP) fusion to its C-terminus and His-tag to its N-terminus, was expressed in prokaryotic cells in soluble form, and highly purified with Ni-Chelating Sepharose Fast Flow column. The purified fusion protein (PEAc1-GFP) efficiently inhibited DNase I activities before polymerization, and activated the myosin Mg-ATPase activities after polymerization. The PEAc1-GFP also polymerized into green fluorescent filamentous structures with a critical concentration of 0.75 uM. These filamentous structures were labeled by TRITC-phalloidin, a specific agent for staining actin microfilaments, and identified as having 9 nm diameters by negative staining. These results indicated that PEAc1 preserved the essential characteristics of actin even with His-tag and GFP fusion, suggesting a promising potential to use GFP fusion protein in obtaining soluble plant actin isoform to analyze its physical and biochemical properties in vitro. The PEAc1-GFP was also expressed in tobacco BY2 cells, which offers a new pathway for further studying its distribution and function in vivo.

Actins↗
PubMed1997

A GFP reporter system to assess gene transfer and expression in human hematopoietic progenitor cells.

Hematopoietic stem cells are widely recognized as attractive targets for gene therapy but current protocols to transduce these cells using recombinant retroviral vectors are inefficient. To evaluate optimization of retroviral transduction of hematopoietic stem cells and stability of gene expression in their progeny, the green fluorescent protein (GFP) was explored as a reporter. We first improved sensitivity of detection > 100-fold over that achieved previously by using a novel retroviral vector (termed MGIN) expressing a high level of an enhanced GFP gene. Primitive human hematopoietic cells bearing the CD34 surface antigen and lacking lineage differentiation markers (CD34+ Lin-) were transduced with the MGIN vector using a clinically applicable supernatant procedure. Under the conditions employed, > 75% of the target cells retained the CD34+ Lin- primitive phenotype after 4-5 days in culture, of those > or = 25% expressed a high level of GFP detectable by both flow cytometric analysis and fluorescence microscopy. When transduced cells were cultured in clonogenic progenitor assays, GFP fluorescence was readily detected in situ, indicating that GFP expression was stable and not detrimental to the differentiative potential of the transduced CD34+ Lin- cells. We conclude that GFP is effective as a vital marker to quantity retrovirus-mediated gene transfer into human hematopoietic and perhaps other types of stem/progenitor cells, and monitor gene expression during their subsequent cell lineage determinations.

Antigens, CD34↗
PubMed2002

Development of a GFP reporter gene for Chlamydomonas reinhardtii chloroplast.

Reporter genes have been successfully used in chloroplasts of higher plants, and high levels of recombinant protein expression have been reported. Reporter genes have also been used in the chloroplast of Chlamydomonas reinhardtii, but in most cases the amounts of protein produced appeared to be very low. We hypothesized that the inability to achieve high levels of recombinant protein expression in the C. reinhardtii chloroplast was due to the codon bias seen in the C. reinhardtii chloroplast genome. To test this hypothesis, we synthesized a gene encoding green fluorescent protein (GFP) de novo, optimizing its codon usage to reflect that of major C. reinhardtii chloroplast-encoded proteins. We monitored the accumulation of GFP in C. reinhardtii chloroplasts transformed with the codon-optimized GFP cassette (GFPct), under the control of the C. reinhardtii rbcL 5'- and 3'-UTRs. We compared this expression with the accumulation of GFP in C. reinhardtii transformed with a non-optimized GFP cassette (GFPncb), also under the control of the rbcL 5'- and 3'-UTRs. We demonstrate that C. reinhardtii chloroplasts transformed with the GFPct cassette accumulate approximately 80-fold more GFP than GFPncb-transformed strains. We further demonstrate that expression from the GFPct cassette, under control of the rbcL 5'- and 3'-UTRs, is sufficiently robust to report differences in protein synthesis based on subtle changes in environmental conditions, showing the utility of the GFPct gene as a reporter of C. reinhardtii chloroplast gene expression.

Amino Acid Sequence↗
PubMed1998

Functional expression of GFP-tagged Kv1.3 and Kv1.4 channels in HEK 293 cells.

Various types of voltage gated potassium channels (Kv) are responsible for setting the resting potential and shaping the membrane potential waveform in the subcellular domains of neurons. In order to visualize the expression behaviour of recombinant Kv channels, we have fused green fluorescent protein (GFP) to the N-terminal of the alpha subunits Kv1.3 and Kv1.4. In transiently transfected HEK 293 cells the GFP-Kv chimeras localize to the plasma membrane. Whole-cell voltage clamp recordings demonstrate that they form functional potassium channels. Kinetic analysis reveals that the gating kinetics of GFP-Kv1.3 are virtually indistinguishable from those displayed by its wild-type correlate. For GFP-Kv1.4 channels we find that their gating is modified in an expected manner. In response to short depolarizing voltage pulses they do not inactivate, indicating that the attached GFP interferes with the fast N-type inactivation mechanism present in wild type Kv1.4 channels. We suggest that GFP tagging of Kv channels might be a useful tool to monitor the spatiotemporal distribution of recombinant potassium channels expressed in living neurons.

Cells, Cultured↗
PubMed2003

Effect of integration of a GFP reporter gene on fitness of Ralstonia eutropha during growth with 2,4-dichlorophenoxyacetic acid.

Green fluorescent proteins (GFPs) are frequently used as marker and reporter systems to assess the fate and activity of microbial strains with the ability to degrade xenobiotic compounds. To evaluate the potential of this tool for tracking herbicide-degrading microorganisms in the environment a promoterless gfp was linked to the tfd C promoter, which is activated during degradation of the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D), and integrated into the chromosome of the 2,4-D-degrading strain Ralstonia eutropha JMP 134. The effects of the inserted gfp gene on the kinetics of 2,4-D degradation by R. eutropha in batch and chemostat culture were compared to those of the wild-type strain. In batch culture with 2,4-D as the only carbon and energy source the maximum specific growth rate of the gfp-marked strain did not differ significantly from the wild type. However, compared to the wild type, the 2,4-D steady-state concentration in 2,4-D-limited chemostat cultures of the gfp-marked strain was higher at all dilution rates tested. The reduced competitiveness of the gfp-marked strain at low substrate concentrations was confirmed in a competition experiment for 2,4-D in continuous culture at a dilution rate of 0.075 h-1. Reproducibly, the gfp-marked strain was displaced by the wild-type strain. The study clearly demonstrates that fitness of constructs cannot be assessed by measuring micro max with selected substrates in batch cultures only but that a thorough kinetic analysis is needed, which also considers slow, carbon-limited growth conditions as they occur in the environment.

2,4-Dichlorophenoxyacetic Acid↗
PubMed2006

Correlated expression of gfp and Bt cry1Ac gene facilitates quantification of transgenic hybridization between Brassicas.

Gene flow from transgenic oilseed rape (BRASSICA NAPUS) might not be avoidable, thus, it is important to detect and quantify hybridization events with its relatives in real time. Data are presented showing the correlation between genetically linked green fluorescent protein (GFP) with BACILLUS THURINGIENSIS (Bt) CRY1AC gene expression in hybrids formed between transgenic B. NAPUS "Westar" and a wild Chinese accession of wild mustard (B. JUNCEA) and hybridization between transgenic B. NAPUS and a conspecific Chinese landrace oilseed rape. Hybrids were obtained either by spontaneous hybridization in the field or by hand-crossing in a greenhouse. In all cases, transgenic hybrids were selected by GFP fluorescence among seedlings originating from seeds harvested from B. JUNCEA and the Chinese oilseed rape plants. Transgenicity was confirmed by PCR detection of transgenes. GFP fluorescence was easily and rapidly detected in the hybrids under greenhouse and field conditions. Results showed that both GFP fluorescence and Bt protein synthesis decreased as either plant or leaf aged, and GFP fluorescence intensity was closely correlated with Bt protein concentration during the entire vegetative lifetime in hybrids. These findings allow the use of GFP fluorescence as an accurate tool to detect gene-flow in time in the field and to conveniently estimate BT CRY1AC expression in hybrids on-the-plant.

Bacillus thuringiensis Toxins↗
PubMed1998

Detection of targeted GFP-Hox gene fusions during mouse embryogenesis.

The ability to use a vital cell marker to study mouse embryogenesis will open new avenues of experimental research. Recently, the use of transgenic mice, containing multiple copies of the jellyfish gene encoding the green fluorescent protein (GFP), has begun to realize this potential. Here, we show that the fluorescent signals produced by single-copy, targeted GFP in-frame fusions with two different murine Hox genes, Hoxa1 and Hoxc13, are readily detectable by using confocal microscopy. Since Hoxa1 is expressed early and Hoxc13 is expressed late in mouse embryogenesis, this study shows that single-copy GFP gene fusions can be used through most of mouse embryogenesis. Previously, targeted lacZ gene fusions have been very useful for analyzing mouse mutants. Use of GFP gene fusions extends the benefits of targeted lacZ gene fusions by providing the additional utility of a vital marker. Our analysis of the Hoxc13(GFPneo) embryos reveals GFP expression in each of the sites expected from analysis of Hoxc13(lacZneo) embryos. Similarly, Hoxa1(GFPneo) expression was detected in all of the sites predicted from RNA in situ analysis. GFP expression in the foregut pocket of Hoxa1(GFPneo) embryos suggests a role for Hoxa1 in foregut-mediated differentiation of the cardiogenic mesoderm.

Aging↗
PubMed1999

Membrane interactions of a constitutively active GFP-Ki-Ras 4B and their role in signaling. Evidence from lateral mobility studies.

Membrane anchorage of Ras proteins in the inner leaflet of the plasma membrane is an important factor in their signaling and oncogenic potential. Despite these important roles, the precise mode of Ras-membrane interactions is not yet understood. It is especially important to characterize these interactions at the surface of intact cells. To investigate Ras-membrane interactions in live cells, we employed studies on the lateral mobility of a constitutively active Ras isoform to characterize its membrane dynamics, and examined the effects of the Ras-displacing antagonist S-trans, trans-farnesylthiosalicylic acid (FTS) (Haklai, R., Gana-Weisz, M., Elad, G., Paz, A., Marciano, D., Egozi, Y., Ben-Baruch, G., and Kloog, Y. (1998) Biochemistry 37, 1306-1314) on these parameters. A green fluorescent protein (GFP) was fused to the N terminus of constitutively active Ki-Ras 4B(12V) to generate GFP-Ki-Ras(12V). When stably expressed in Rat-1 cells, this protein was preferentially localized to the plasma membrane and displayed transforming activity. The lateral mobility studies demonstrated that GFP-Ki-Ras(12V) undergoes fast lateral diffusion at the plasma membrane, rather than exchange between membrane-bound and unbound states. Treatment of the cells with FTS had a biphasic effect on GFP-Ki-Ras(12V) lateral mobility. At the initial phase, the lateral diffusion rate of GFP-Ki-Ras(12V) was elevated, suggesting that it is released from some constraints on its lateral mobility. This was followed by dislodgment of the protein into the cytoplasm, and a reduction in the diffusion rate of the fraction of GFP-Ki-Ras(12V) that remained associated with the plasma membrane. Control experiments with other S-prenyl analogs showed that these effects are specific for FTS. These results have implications for the interactions of Ki-Ras with specific membrane anchorage domains or sites.

Animals↗
PubMed2002

Col1a1-GFP transgene expression in developing incisors.

Previous studies have shown that terminal differentiation of odontoblasts is accompanied by dramatic increases in type I collagen synthesis. Recently transgenic mice in which green fluorescent protein (GFP) expression is under the control of the rat 3.6 (pOBCol3.6GFPtpz) and 2.3 (pOBCol2.3GFPemd) Col1a1 promoter fragments were generated. Our analysis of these GFP-expressing transgenic mice shows that the 2.3-kb promoter fragment directs strong expression of GFP only to bones and teeth, whereas the 3.6-kb fragment of promoter directs strong expression of GFP in bone and tooth, as well as in other type I collagen producing tissues. Our observations of incisors in these transgenic mice show high levels of GFP expression in functional odontoblasts and in differentiated osteoblasts. These observations show that expression of GFP reporter genes closely follow the patterns of expression of alpha 1(I) collagen in various tissues including odontoblasts.

Aging↗
PubMed2006

Faithful expression of GFP from the PLCbeta2 promoter in a functional class of taste receptor cells.

Phospholipase C-type beta2 (PLCbeta2) is expressed in a subset of cells within mammalian taste buds. This enzyme is involved in the transduction of sweet, bitter, and umami stimuli and thus is believed to be a marker for gustatory sensory receptor cells. We have developed transgenic mice expressing green fluorescent protein (GFP) under the control of the PLCbeta2 promoter to enable one to identify these cells and record their physiological activity in living preparations. Expression of GFP (especially in lines with more than one copy integrated) is strong enough to be detected in intact tissue preparations using epifluorescence microscopy. By immunohistochemistry, we confirmed that the overwhelming majority of cells expressing GFP are those that endogenously express PLCbeta2. Expression of the GFP transgene in circumvallate papillae occurs at about the same time during development as endogenous PLCbeta2 expression. When loaded with a calcium-sensitive dye in situ, GFP-positive taste cells produce typical Ca2+ responses to a taste stimulus, the bitter compound cycloheximide. These PLCbeta2 promoter-GFP transgenic lines promise to be useful for studying taste transduction, sensory signal processing, and taste bud development.

Animals↗
PubMed2001

Assessment of GFP fluorescence in cells of Streptococcus gordonii under conditions of low pH and low oxygen concentration.

Use of green fluorescent protein (GFP) as a molecular reporter is restricted by several environmental factors, such as its requirement for oxygen in the development of the fluorophore, and its poor fluorescence at low pH. There are conflicting data on these limitations, however, and systematic studies to assess the importance of these factors for growing bacterial cultures are lacking. In the present study, homogeneous expression of the gfpmut3* gene directed by a synthetic constitutive lactococcal promoter was demonstrated in batch cultures and in biofilms of Streptococcus gordonii DL1. A lower limit of oxygen concentration for maturation of the GFP fluorophore was determined: fluorescence was emitted at 0.1 p.p.m. dissolved oxygen (in conventionally prepared anaerobic media lacking reducing agents), whereas no fluorescence was detected in the presence of 0.025 p.p.m. dissolved oxygen (obtained by addition of L-cysteine as reducing agent). When an anaerobically grown (non-fluorescent) >50 microm thick biofilm was shifted to aerobic conditions, fluorescence could be detected within 4 min, reaching a maximum over the next 16 min. It was not possible to detect any fluorescence gradients (lateral or vertical) within the >50 microm thick biofilm, and fluorescence development after the shift to aerobic conditions occurred throughout the biofilm (even at the substratum). This suggests that oxygen gradients, which might result in reduced GFP fluorescence, did not exist in the >50 microm thick biofilm of this organism. Production of lactic acid and the subsequent acidification in batch cultures of S. gordonii DL1 led to a decrease in fluorescence intensity. However, severe pH reduction was prevented when the bacterium was grown as a biofilm in a flowcell, and a homogeneous distribution of a strong fluorescence signal was observed. These findings show that GFP can be applied to studies of oxygen-tolerant anaerobic bacteria, that densely packed, flowcell-grown biofilms of S. gordonii do not develop oxygen gradients inhibitory to GFP fluorescence development, and that the often transient nature of GFP fluorescence in acid-producing bacteria can be overcome in flowcells, probably by the elimination of metabolic by-product accumulation.

Anaerobiosis↗
PubMed2004

Propagation of gammaPKC translocation along the dendrites of Purkinje cell in gammaPKC-GFP transgenic mice.

To elucidate spatial and temporal profiles of the protein kinase C (PKC) activation in relation to neuronal functions including synaptic plasticity, we tried to detect PKC translocation in living brain slices. We first developed brain region-specific and inducible gammaPKC-GFP transgenic mice using a tetracycline (tet)-regulated system. In the transgenic mice, the expression of gammaPKC-GFP was region-specifically regulated by the promoter and abolished by the administration of doxycycline. Cerebellar slices from the mice were utilized for intracellular recording and fluorescence imaging of gammaPKC-GFP in Purkinje cells. GFP fluorescence was uniformly distributed from soma to dendritic arbor. When mGluR agonists were applied, the intensity was transiently increased at the edge of the dendrite and concomitantly decreased in the cytoplasm, indicating that gammaPKC translocated to the plasma membrane. This transient change in the pattern of GFP fluorescence simultaneously occurred throughout the Purkinje cell dendrites by agonist stimulation. Translocation of gammaPKC-GFP was also induced by electrical stimulation of parallel fibres. However, the event was not restricted at the distal dendrites, propagated forwardly along the dendritic tree and reached to the proximal trunk close to the soma. Time course of the propagation was slower than the electrical signal and Ca(2+) waves and faster than conveying molecules through microtubules. The present results indicate that PKC signals activated locally by parallel fibre input could propagate to the soma through dendrites in living Purkinje neurones. The findings may provide us with a new insight for understanding molecular mechanisms of the synaptic plasticity including cerebellar long-term depression.

Animals↗
PubMed2004

Viroid-induced RNA silencing of GFP-viroid fusion transgenes does not induce extensive spreading of methylation or transitive silencing.

Viroid infection is associated with the production of short interfering RNAs (siRNAs), a hallmark of post-transcriptional gene silencing (PTGS). However, viroid RNAs autonomously replicating in the nucleus have not been shown to trigger the degradation of homologous RNA in the cytoplasm. To investigate the potential of viroids for the induction of gene silencing, non-infectious fragments of potato spindle tuber viroid (PSTVd) cDNA were transcriptionally fused to the 3' end of the green fluorescent protein (GFP)-coding region. Introduction of such constructs into tobacco plants resulted in stable transgene expression. Upon PSTVd infection, transgene expression was suppressed and partial de novo methylation of the transgene was observed. PSTVd-specific siRNA was detected but none was found corresponding to the gfp gene. Methylation was restricted almost entirely to the PSTVd-specific part of the transgene. Neither a gfp transgene construct lacking viroid-specific elements was silenced nor was de novo methylation detected, when it was introduced into the genetic background of the PSTVd-infected plant lines containing silenced GFP:PSTVd transgenes. The absence of gfp-specific siRNAs and of significant methylation within the gfp-coding region demonstrated that neither silencing nor DNA methylation spread from the initiator region into adjacent 5' regions.

Base Sequence↗
PubMed2002

Construction and characterization of a proU-gfp transcriptional fusion that measures water availability in a microbial habitat.

We constructed and characterized a transcriptional fusion that measures the availability of water to a bacterial cell. This fusion between the proU promoter from Escherichia coli and the reporter gene gfp was introduced into strains of E. coli, Pantoea agglomerans, and Pseudomonas syringae. The proU-gfp fusion in these bacterial biosensor strains responded in a quantitative manner to water deprivation caused by the presence of NaCl, Na(2)SO(4), KCl, or polyethylene glycol (molecular weight, 8000). The fusion was induced to a detectable level by NaCl concentrations of as low as 10 mM in all three bacterial species. Water deprivation induced proU-gfp expression in both planktonic and surface-associated cells; however, it induced a higher level of expression in the surface-associated cells. Following the introduction of P. agglomerans biosensor cells onto bean leaves, the cells detected a significant decrease in water availability within only 5 min. After 30 min, the populations were exposed, on average, to a water potential equivalent to that imposed by approximately 55 mM NaCl. These results demonstrate the effectiveness of a proU-gfp-based biosensor for evaluating water availability on leaves. Furthermore, the inducibility of proU-gfp in multiple bacterial species illustrates the potential for tailoring proU-gfp-based biosensors to specific habitats.

Amino Acid Transport Systems↗