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Hygromycin B

Hygromycin B is an antibiotic produced by the bacterium Streptomyces hygroscopicus.
It is commonly used as a selective marker in genetic engineering and cell culture applications.
Hygromycin B works by inhibiting protein synthesis in eukaryotic cells, making it an effective tool for identifying and isolating cells that have been successfully transfected or transformed.
This MeSH term provides a concise overview of the key properties and applications of Hygromycin B, a widely used reagent in molecular biology and cell biology researcg.

Most cited protocols related to «Hygromycin B»

1
Robust Knock-in Mouse Line Generation
Cited 721 times
Targeting constructs were generated using a combined gene synthesis (GenScript Corp.) and molecular cloning approach. Briefly, to target the Rosa26 locus, a cassette containing the following components was constructed: FRT – LoxP – Stop codons – 3x SV40 polyA – LoxP – EYFP – WPRE – bGH polyA – AttB – PGK promoter – FRT – Neo – PGK polyA – AttP. For most targeting vectors, this cassette was cloned into a Rosa-CAG targeting vector3 (link), downstream of the CAG promoter and upstream of the 3′ arm, to generate the final EYFP targeting vector. Unique restriction sites flanking the EYFP gene were used to replace EYFP with alternative reporter genes. For the Ai2 vector, which lacks the WPRE, the CAG promoter was inserted between the first FRT and LoxP sites, and the cassette was cloned immediately downstream of the 5′ homology arm. The final targeting vectors contained 5′ and 3′ homology arms of 1.1 kb and 4.3 kb, as well as a PGK-DTA cassette for negative selection. Targeting constructs for knock-in Cre lines inserted into other gene loci were constructed in similar ways.
The targeting vectors were linearized and transfected into the 129/B6 F1 hybrid ES cell line G442 (link) using an Amaxa electroporator. G418-resistant ES clones were screened by Southern blot analysis of HindIII digested DNA, which was probed with a 1.1 kb genomic fragment from immediately upstream of the 5′ arm. We observed a recombination rate of about 25% for the four constructs. Positive ES clones were injected into C57BL/6J blastocysts to obtain chimeric mice following standard procedures. Both ES cell transfections and blastocyst injections were performed by the University of Washington Transgenic Resources Program. Due to the robustness of the G4 cells, high-percentage chimeras and high rates of germline transmission were routinely obtained. Chimeric mice were bred with either C57BL/6J mice to obtain germline transmission or various Cre-driver lines for direct characterization.
An Ai9 ES cell clone with strong germline transmission potency was used in subsequent transfections for the Flp-mediated exchange strategy outlined in Supplementary Figure 4 online. Ai9 ES cells were co-transfected using a Bio-Rad electroporator with 100 μg of pCAGGS-FLPe (Open Biosystems) and 40 μg of an incoming replacement vector. After 8 to 10 days of Hygromycin B selection, surviving colonies that also appeared green by fluorescence microscopy were picked and screened by PCR using primer sets designed to confirm a correct insertion of the incoming vector at the 5′ and 3′ FRT recombinase sites.
Madisen L., Zwingman T.A., Sunkin S.M., Oh S.W., Zariwala H.A., Gu H., Ng L.L., Palmiter R.D., Hawrylycz M.J., Jones A.R., Lein E.S, & Zeng H. (2009). A robust and high-throughput Cre reporting and characterization system for the whole mouse brain. Nature neuroscience, 13(1), 133-140.
Publication 2009
Anabolism Animals, Transgenic antibiotic G 418 Blastocyst Cells Chimera Clone Cells Cloning Vectors Codon, Terminator Embryonic Stem Cells Genes Genes, Reporter Genetic Loci Genome Germ Cells Germ Line Hybrid Cells Hygromycin B Mice, Inbred C57BL Microscopy, Fluorescence Mus N-fluoresceinylphosphatidylethanolamine Oligonucleotide Primers Poly A Recombinase Recombination, Genetic Rosa Simian virus 40 Southern Blotting Transfection Transmission, Communicable Disease
2
Robust Knock-in Mouse Line Generation
Cited 686 times
Targeting constructs were generated using a combined gene synthesis (GenScript Corp.) and molecular cloning approach. Briefly, to target the Rosa26 locus, a cassette containing the following components was constructed: FRT – LoxP – Stop codons – 3x SV40 polyA – LoxP – EYFP – WPRE – bGH polyA – AttB – PGK promoter – FRT – Neo – PGK polyA – AttP. For most targeting vectors, this cassette was cloned into a Rosa-CAG targeting vector3 (link), downstream of the CAG promoter and upstream of the 3′ arm, to generate the final EYFP targeting vector. Unique restriction sites flanking the EYFP gene were used to replace EYFP with alternative reporter genes. For the Ai2 vector, which lacks the WPRE, the CAG promoter was inserted between the first FRT and LoxP sites, and the cassette was cloned immediately downstream of the 5′ homology arm. The final targeting vectors contained 5′ and 3′ homology arms of 1.1 kb and 4.3 kb, as well as a PGK-DTA cassette for negative selection. Targeting constructs for knock-in Cre lines inserted into other gene loci were constructed in similar ways.
The targeting vectors were linearized and transfected into the 129/B6 F1 hybrid ES cell line G442 (link) using an Amaxa electroporator. G418-resistant ES clones were screened by Southern blot analysis of HindIII digested DNA, which was probed with a 1.1 kb genomic fragment from immediately upstream of the 5′ arm. We observed a recombination rate of about 25% for the four constructs. Positive ES clones were injected into C57BL/6J blastocysts to obtain chimeric mice following standard procedures. Both ES cell transfections and blastocyst injections were performed by the University of Washington Transgenic Resources Program. Due to the robustness of the G4 cells, high-percentage chimeras and high rates of germline transmission were routinely obtained. Chimeric mice were bred with either C57BL/6J mice to obtain germline transmission or various Cre-driver lines for direct characterization.
An Ai9 ES cell clone with strong germline transmission potency was used in subsequent transfections for the Flp-mediated exchange strategy outlined in Supplementary Figure 4 online. Ai9 ES cells were co-transfected using a Bio-Rad electroporator with 100 μg of pCAGGS-FLPe (Open Biosystems) and 40 μg of an incoming replacement vector. After 8 to 10 days of Hygromycin B selection, surviving colonies that also appeared green by fluorescence microscopy were picked and screened by PCR using primer sets designed to confirm a correct insertion of the incoming vector at the 5′ and 3′ FRT recombinase sites.
Madisen L., Zwingman T.A., Sunkin S.M., Oh S.W., Zariwala H.A., Gu H., Ng L.L., Palmiter R.D., Hawrylycz M.J., Jones A.R., Lein E.S, & Zeng H. (2009). A robust and high-throughput Cre reporting and characterization system for the whole mouse brain. Nature neuroscience, 13(1), 133-140.
Publication 2009
Anabolism Animals, Transgenic antibiotic G 418 Blastocyst Cells Chimera Clone Cells Cloning Vectors Codon, Terminator Embryonic Stem Cells Genes Genes, Reporter Genetic Loci Genome Germ Cells Germ Line Hybrid Cells Hygromycin B Mice, Inbred C57BL Microscopy, Fluorescence Mus N-fluoresceinylphosphatidylethanolamine Oligonucleotide Primers Poly A Recombinase Recombination, Genetic Rosa Simian virus 40 Southern Blotting Transfection Transmission, Communicable Disease
3
HTLA Cell-Based Luciferase Assay
Cited 55 times
HTLA cells, (an HEK293 cell line stably expressing a tTA-dependent luciferase reporter and a β-arrestin2-TEV fusion gene) were a gift from the lab of Richard Axel, and were maintained in DMEM supplemented with 10% FBS, 2 μg/ml puromycin and 100 μg/ml hygromycin B in a humidified atmosphere at 37°C in 5% CO2. For transfection, cells were plated at 9 to 10 × 106 cells per 150 mm cell culture dish (day 1). The following day (day 2), cells were transfected using the calcium phosphate method. On day 3, transfected cells were transferred at 15,000 to 20,000 cells per well in 40 μl of medium into poly-L-lysine coated and rinsed 384-well white clear-bottom cell culture plates (Greiner Bio-one). On day 4, 3.5x drug stimulation solutions were prepared in filter-sterilized assay buffer, which consisted of 20 mM HEPES and 1x HBSS at pH 7.4, and 20 μl added to each well. On day 5, medium and drug solutions were removed from the wells (by aspiration or shaking), and 20 μl per well of Bright-Glo solution (Promega) diluted 20-fold in assay buffer were added to each well. After incubation for 15 to 20 minutes at room temperature, luminescence was counted in a Trilux luminescence counter. Results in the form of RLU (relative luminescence units) were exported into Excel spreadsheets, and Graphpad Prism was used for analysis of data. To measure constitutive activity, no ligand was added on day 4.
Kroeze W.K., Sassano M.F., Huang X.P., Lansu K., McCorvy J.D., Giguere P.M., Sciaky N, & Roth B.L. (2015). PRESTO-TANGO: an open-source resource for interrogation of the druggable human GPCR-ome. Nature structural & molecular biology, 22(5), 362-369.
Publication 2015
Atmosphere beta-Arrestin 1 Biological Assay Buffers Calcium Phosphates Cell Culture Techniques Cells Genes, vif HEK293 Cells Hemoglobin, Sickle HEPES Hygromycin B Hyperostosis, Diffuse Idiopathic Skeletal Leukocytes Ligands Luciferases Luminescence Lysine Poly A prisma Promega Puromycin Transfection
4
Yeast Cultivation and Transformation Protocols
Cited 55 times
All experiments were carried out in the DF5 strain background (Finley et al., 1987 (link)). All strains used are listed in Table S2 (see Supporting information). Standard procedures were followed for yeast cultivation and transformation (Sherman, 1991 (link)). For use of the CUP1 promoter, 100 µM CuSO4 was added to the growth medium. Geneticin was used at 200 µg/ml (for kanMX4 selection); hygromycin B at 300 µg/ml (for hphNT1); and nourseothricin at 100 µg/ml (for natNT2). Auxin was used at 1 mm unless otherwise indicated. TIR1 strains were created by integration of pNHK53 (encoding OsTIR1 under control of the ADH1 promoter) into the URA3 locus (Nishimura et al., 2009 (link)). All strains carrying a deletion or epitope-tagged allele of RAD53 were constructed in an sml1Δ background. Gene deletions and tags were introduced by means of PCR-generated cassettes (Longtine et al., 1998 (link)), using the primers listed in Table S1 (see Supporting information).
Morawska M, & Ulrich H.D. (2013). An expanded tool kit for the auxin-inducible degron system in budding yeast. Yeast (Chichester, England), 30(9), 341-351.
Publication 2013
Alleles Auxins Culture Media Deletion Mutation Epitopes Gene Deletion Geneticin Hygromycin B Nourseothricin Oligonucleotide Primers Saccharomyces cerevisiae Strains
5
Arabidopsis Transformation and Reporter Gene Assays
Cited 45 times
Transformations of Arabidopsis thaliana were performed by the floral dip method [2 (link)] using Agrobacterium tumefaciens strain GV3101 [15 (link)]. The following binary plasmids were used: pBINPLUS [5 (link)] which confers kanamycin resistance via the nptII gene; pSKI015 [7 (link)] which confers phosphinothricin resistance via the bar gene; and pBIG-HYG [8 (link)] which confers hygromycin B resistance via the hpt gene. The reporter genes uidA with added intron [6 (link)] and mGFP4 [4 (link)] were inserted into the binary vector pBINPLUS under the control of 707 bp of the 5' upstream sequence of the carbonic anhydrase 1 (CA1) gene (At3g01500). The following non-transformed Arabidopsis seeds, obtained from the Nottingham Arabidopsis stock centre (NASC), were used for transformations and as controls; Col-0 (NASC N1092), Col-2 (NASC N907), Col-7 (NASC N3731), Ws (NASC N1601) and Ler-0 (NASC NW20). Previously characterised transgenic lines were used as positive controls; for phosphinothricin-resistance NASC accessions N21504, N21443, N21461, N21821, N21824 and N850573 were used, and for hygromycin-resistance 5 lines transformed with pBIG-HYG containing mGFP4 (K. Parsley unpublished data) were used.
Harrison S.J., Mott E.K., Parsley K., Aspinall S., Gray J.C, & Cottage A. (2006). A rapid and robust method of identifying transformed Arabidopsis thaliana seedlings following floral dip transformation. Plant Methods, 2, 19.
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Publication 2006
Agrobacterium tumefaciens Animals, Transgenic Arabidopsis Arabidopsis thalianas Cloning Vectors Dehydratase, Carbonate Gene Expression Regulation Genes Genes, Reporter hygromycin A Hygromycin B Introns Kanamycin Resistance Parsley phosphinothricin Plant Embryos Plasmids Strains

Most recents protocols related to «Hygromycin B»

1
Quantitative ER-Mitochondria Interaction Assay
Mouse monoclonal anti ACTIN, Sigma-Aldrich, A5441; Mouse monoclonal anti c-myc, Sigma-Aldrich, M4439; Rabbit polyclonal anti Calnexin, Enzo, ADI-SPA-865-F; Mouse monoclonal anti CYTC, BD Bioscience, 556433; Rabbit DyLight 680, Thermo Fisher Scientific, 35569; Mouse DyLight 680 Thermo Fisher Scientific, 35519; Mouse DyLight 800 Thermo Fisher Scientific, 35521; Rabbit DyLight 800 Thermo Fisher Scientific, 35571; Mouse monoclonal anti eIF2α, Cell Signaling, 2103S; Rabbit polyclonal anti-E-Syt1, Sigma-Aldrich, HPA016858; Rabbit polyclonal anti-GFP, Cell Signaling, 2555S; Mouse monoclonal anti-GFP,Life technologies, A11122; HRP Mouse Bioké, Cell Signaling, 7076; HRP Rabbit Bioké, Cell Signaling, 7074; Mouse monoclonal anti IP3R3, BD Bioscience, 610312; Rabbit polyclonal anti-PERK, Cell signaling, 3192S; Rabbit polyclonal anti PERK, Cell signaling, 5683S; Rabbit monoclonal anti Phospho-eIF2α (Ser51), Cell signaling, 3597S; Rabbit polyclonal anti PDI Genetex, GTX30716; Mouse monoclonal anti PSD, Santa Cruz, sc-390070; Rabbit polyclonal anti, PSS1 (B-5), Santa Cruz, sc-515376; Rabbit polyclonal anti PSS2, Sigma-Aldrich, SAB1303408; Rabbit polyclonal VDAC1, Cell Signaling, 4866S; Rabbit polyclonal VDAC1, Abcam, ab15895; Veriblot antibody Abcam, ab131366.
The reagents used were: Antimycin A, Sigma-Aldrich, A8674; Calcium Chloride dihydrate, Sigma-Aldrich, C3881; CHAPS hydrate, Sigma-Aldrich, C3023; Conjugated GFP antibody beads, Laboratory of Chris Ulens; D-Galactose, Sigma-Aldrich, G0750; D-glucose, Sigma-Aldrich, G7021-1KG; DAPI, Thermo Fisher Scientific, 62248; Dulbecco’s Modified Eagle’s Medium - high glucose, Sigma-Aldrich, D0422; EGTA, AppliChem, A0878; FCCP, Sigma-Aldrich, C2920; Gibco DMEM/F-12, Thermo Fisher Scientific, 11320074; Glucose, Agilent Seahorse, 103577; Glutamine, Sigma-Aldrich, G7513; Glutamine, Agilent Seahorse, 103579; GSK PERK Inhibitor, Toronto Research Company, G797800; Hygromycin B, Invivogen, ant-hg-1; Lipofectamine 2000 Transfection Reagent, Thermo Fisher Scientific, 11668019; MitoTracker FarRed, Thermo Fisher Scientific, M22426; NBD-PS, Avanti Polar Lipids, 810194C; SE Cell Line 4D-Nucleofector X Kit L, V4XC-1024; Oligomycin, Sigma-Aldrich, 75351; Penicillin and streptomycin, Sigma-Aldrich, P0781; Percoll, Sigma-Aldrich, P1644; Pierce ECL Western Blotting Substrate, Thermo Fisher Scientific, 32106X4; Pierce Protein A/G Magnetic Beads, Thermo Fisher Scientific, 88802; Pierce Protease Inhibitor Tablets, EDTA-free, Thermo Fisher Scientific, 88266; Potassium Chloride, Janssen Chimica, 7447407; Protease inhibitor, Thermo Fisher Scientific, A32953; Puromycin, Thermo Fisher Scientific, A11138-03; Protein A/G PLUS-Agarose, Santa Cruz, sc-2003; XF DMEM pH7 7.4, Agilent Seahorse, 103575; Sodium Chloride, Sigma-Aldrich, A0431796; Sodium Pyruvate Solution, Agilent Seahorse, 103578; Sucrose, Acros, A0333146; Thapsigargin, Enzo Life Sciences, BML-PE180; TransIT-X2 Dynamic Delivery System, Mirus Bio, MIR 6000; Tris base, Sigma-Aldrich, 77861; Triton, Sigma-Aldrich, T9234; Tween, Sigma Aldrich, P4780.
Sassano M.L., van Vliet A.R., Vervoort E., Van Eygen S., Van den Haute C., Pavie B., Roels J., Swinnen J.V., Spinazzi M., Moens L., Casteels K., Meyts I., Pinton P., Marchi S., Rochin L., Giordano F., Felipe-Abrio B, & Agostinis P. (2023). PERK recruits E-Syt1 at ER–mitochondria contacts for mitochondrial lipid transport and respiration. The Journal of Cell Biology, 222(3), e202206008.
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Publication 2023
3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate Actins Antimycin A Calcium Chloride Dihydrate Calnexin Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone DAPI Eagle Edetic Acid Egtazic Acid G-substrate Galactose Glucose Glutamine Hygromycin B Immunoglobulins L Cells Lipids lipofectamine 2000 Mus N-(7-nitro-2,1,3-benzoxadiazol-4-yl)phosphatidylserine Obstetric Delivery Oligomycins Peeling Skin Syndrome Peeling skin syndrome, acral type Penicillins Percoll Potassium Chloride Protease Inhibitors Puromycin Pyruvate Rabbits Seahorses Sepharose Sodium Sodium Chloride Staphylococcal Protein A Streptomycin Sucrose SYT1 protein, human Thapsigargin Transfection Tromethamine Tweens VDAC1 protein, human
2
Cell Culture and Transfection for Live-Cell Imaging
U2OS, COS-7, Vero-E6, HeLa, Caco-2, and Swiss 3T3 cells were cultured in DMEM supplement with 10% FBS and 50 µg/ml penicillin/streptomycin at 37°C and 5% CO2. U2OS Flp-In T-Rex cells were cultured in DMEM supplement with 10% FBS, 50 µg/ml penicillin/streptomycin, and 15 µg/ml blastidicin. U2OS, Vero-E6, and HeLa cells were purchased from ATCC; Swiss 3T3 and COS-7 cells were a kind gift from Anna Akhmanova; U2OS Flp-In T-Rex cells were a kind gift from Prof. Alessandro Sartori (Institute of Molecular Cancer Research, University of Zurich, Zurich, Switzerland); and Caco-2 cells were a kind gift from Sven van Ijzendoorn (University Medical Center Groningen, Groningen, Netherlands). Cells were confirmed to be free of mycoplasma. Cells were plated on 18-mm coverslips (immunolabeling experiments) or 25-mm coverslips (live-cell experiments) on the day of or 1 or 2 d before transient transfection. Cells were transiently transfected using Fugene6 transfection reagent (Promega) according to the manufacturer’s protocol. After 1 d of transfection, cells were used for live-cell imaging, subjected to treatment (drugs/serum starvation), or fixated. For all experiments that involved the expression of mCherry-tubulin, cells were used for live-cell imaging after 2 d of transfection. The isogenic U2OS Flp-In cell line that upon doxycycline-induction stably expresses hKif5b(1–560)G234A-mNeongreen-mNeongreen was derived from the U2OS Flp-In cell line by transfection with the pCDN5/FRT/TO vector (Invitrogen) and pOG44 vector (Invitrogen). The U2OS Flp-In hKif5b(1–560)G234A-L-mNeongreen-L-mNeongreen cell line was cultured in DMEM supplemented with 10% FBS, 50 µg/ml penicillin/streptomycin, 15 µg/ml blastidicin, and 0.5 mg/ml hygromycin B (ant-hg-1, InvivoGen). To induce expression of hKif5b(1-560)G234A-L-mNeongreen-L-mNeongreen, 10 ng doxycycline (ab141091; Abcam) was added to the cells 16–24 h before imaging. U2OS Flp-In hKif5b(1-560)G234A-L-mNeongreen-L-mNeongreen cells were plated on 25-mm coverslips ≥2 d before imaging.
Jansen K.I., Iwanski M.K., Burute M, & Kapitein L.C. (2023). A live-cell marker to visualize the dynamics of stable microtubules throughout the cell cycle. The Journal of Cell Biology, 222(5), e202106105.
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Publication 2023
Caco-2 Cells Cell Lines Cells Cloning Vectors COS-7 Cells Doxycycline HeLa Cells Hygromycin B Malignant Neoplasms Mycoplasma Penicillins Pharmaceutical Preparations Promega Serum Streptomycin Swiss 3T3 Cells T-Lymphocyte Transfection Transients Tubulin
3
Calcium Mobilization Assay for Opioid Receptors
Stock solutions (10 mM) of the peptides were prepared in 5% DMSO in bidistilled water and kept at −20°C Serial dilutions were carried out in HBSS/HEPES buffer (20 mM, containing 0.02% bovine serum albumin fraction V). Calcium mobilization assay was performed using the same method as previously described (Camarda and Calo, 2013 (link)). CHO cells with stable co-expression of human mu or kappa receptors and the C-terminally modified Gαqi5 and CHO cells with co-expression of the delta-opioid receptor and the GαqG66Di5 protein were used. Dulbecco’s MEM/HAMS F12 (1:1) medium supplemented with 10% fetal bovine serum, penicillin (100 IU/mL), streptomycin (100 μg/mL), L-glutammine (2 mM), fungizone (1 μg/mL), geneticin (G418; 200 μg/mL) and hygromycin B (100 μg/mL) was used for cell culture. Cells were seeded at a density of 50,000 cells/well into 96-well black, clear-bottom plates and kept in the incubator at 37°C in 5% CO2/humidified air. After 24 h the cell growth medium was aspired and loading medium, supplemented with probenecid (2.5 mM), calcium sensitive fluorescent dye Fluo-4 AM (3 µM), pluronic acid (0.01%) and HEPES (20 mM), was added. Then the plates were placed in the incubator again. After 30 min the loading solution was aspirated and 100 µL/well of assay buffer (HBSS supplemented with 20 mM HEPES, 2.5 mM probenecid, and 500 µM Brilliant Black) was added. Next, both plates (cell culture and compound plate) were placed in the FlexStation II reader (Molecular Device, Union City, CA 94587, United States), the on-line additions were carried out in a volume of 50 µL/well and the fluorescence changes were measured. Ligand efficacies, expressed as the intrinsic activity (α), were calculated as the Emax ratio of the tested compound and the standard agonist. At least three independent experiments for each assay were carried out in duplicate.
Curve fittings were performed using Graph Pad PRISM 5.0 (GraphPad Software Inc., San Diego, United States). Data have been statistically analyzed with one way ANOVA followed by the Dunnett’s test for multiple comparisons; p values < 0.05 were considered significant.
Piekielna-Ciesielska J., Malfacini D., Djeujo F.M., Marconato C., Wtorek K., Calo’ G, & Janecka A. (2023). Functional selectivity of EM-2 analogs at the mu-opioid receptor. Frontiers in Pharmacology, 14, 1133961.
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Publication 2023
Acids antibiotic G 418 ATF7IP protein, human Biological Assay brilliant black Buffers Calcium Cell Culture Techniques Cells CHO Cells Fetal Bovine Serum Fluo 4 Fluorescence Fluorescent Dyes Fungizone Geneticin Hemoglobin, Sickle HEPES Homo sapiens Hygromycin B Ligands Medical Devices neuro-oncological ventral antigen 2, human Penicillins Peptides Pluronics prisma Probenecid Proteins Receptors, Cell Surface Receptors, Opioid, delta Receptors, Opioid, kappa Receptors, Opioid, mu Serial C Serum Albumin, Bovine Streptomycin Sulfoxide, Dimethyl Technique, Dilution
4
Auxin-Induced Degradation of DPY30 and RBBP5
For the generation of the auxin-inducible degradation system for DPY30 sgRNA targeting the stop-codon region were cloned into eSpCas9(1.1)-T2A-eGFP. Left and right homology arms, as well as the mAID-T2A-BFP middle part were ligated into a modified pUC19 vector backbone (a gift from S. Pollard) using the In-Fusion cloning kit (Takara, 638910). mES cells were co-transfected with sgRNA- and donor-vector using Lipofectamine 3000 and sorted 48 h later for GFP/BFP-double-positive cells. Homozygous clones were then transfected with pPB-hygro-OsTIR1-P2A-mCherry and pBase plasmids and selected with 100 µg ml−1 hygromycin B (Thermo Fisher Scientific, 10687010). For the generation of the endogenous dTAG-inducible degradation system for RBBP5, sgRNA targeting the stop codon region were co-transfected into the cells and contained the following elements: left and right homology arms, as well as FKBP12(F36V), 2× HA tags, P2A and a neomycin-resistance gene. The transfected cells were selected with 100 µg ml−1 Geneticin selective antibiotic (G418 Sulfate) (Thermo Fisher Scientific, 10131027), single-cell sorted to obtain clonal cell lines and screened for correct biallelic integration. All homozygous insertions and knock-ins were confirmed by Sanger sequencing and western blotting. A list of the oligos and the sequences of the sgRNAs is provided in Supplementary Table 1.
Wang H., Fan Z., Shliaha P.V., Miele M., Hendrickson R.C., Jiang X, & Helin K. (2023). H3K4me3 regulates RNA polymerase II promoter-proximal pause-release. Nature, 615(7951), 339-348.
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Publication 2023
2',5'-oligoadenylate antibiotic G 418 Antibiotics Arm, Upper Auxins Cell Lines Cells Clone Cells Cloning Vectors Codon, Terminator Genes Geneticin Homozygote Hygromycin B Insertion Mutation Lipofectamine Neomycin Plasmids Sulfates, Inorganic Tacrolimus Binding Protein 1A Tissue Donors Vertebral Column
5
Auxin-Induced Degradation of DPY30 and RBBP5
For the generation of the auxin-inducible degradation system for DPY30 sgRNA targeting the stop-codon region were cloned into eSpCas9(1.1)-T2A-eGFP. Left and right homology arms, as well as the mAID-T2A-BFP middle part were ligated into a modified pUC19 vector backbone (a gift from S. Pollard) using the In-Fusion cloning kit (Takara, 638910). mES cells were co-transfected with sgRNA- and donor-vector using Lipofectamine 3000 and sorted 48 h later for GFP/BFP-double-positive cells. Homozygous clones were then transfected with pPB-hygro-OsTIR1-P2A-mCherry and pBase plasmids and selected with 100 µg ml−1 hygromycin B (Thermo Fisher Scientific, 10687010). For the generation of the endogenous dTAG-inducible degradation system for RBBP5, sgRNA targeting the stop codon region were co-transfected into the cells and contained the following elements: left and right homology arms, as well as FKBP12(F36V), 2× HA tags, P2A and a neomycin-resistance gene. The transfected cells were selected with 100 µg ml−1 Geneticin selective antibiotic (G418 Sulfate) (Thermo Fisher Scientific, 10131027), single-cell sorted to obtain clonal cell lines and screened for correct biallelic integration. All homozygous insertions and knock-ins were confirmed by Sanger sequencing and western blotting. A list of the oligos and the sequences of the sgRNAs is provided in Supplementary Table 1.
Wang H., Fan Z., Shliaha P.V., Miele M., Hendrickson R.C., Jiang X, & Helin K. (2023). H3K4me3 regulates RNA polymerase II promoter-proximal pause-release. Nature, 615(7951), 339-348.
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Publication 2023
2',5'-oligoadenylate antibiotic G 418 Antibiotics Arm, Upper Auxins Cell Lines Cells Clone Cells Cloning Vectors Codon, Terminator Genes Geneticin Homozygote Hygromycin B Insertion Mutation Lipofectamine Neomycin Plasmids Sulfates, Inorganic Tacrolimus Binding Protein 1A Tissue Donors Vertebral Column

Top products related to «Hygromycin B»

1
Hygromycin b by Thermo Fisher Scientific
Sourced in United States, United Kingdom, France, Germany, China, Japan, Switzerland, Italy, Austria, Australia
Hygromycin B is a laboratory reagent used for antibiotic selection during genetic engineering and cell culture experiments. It is a broad-spectrum aminoglycoside antibiotic that inhibits protein synthesis in eukaryotic cells. Hygromycin B is commonly used as a selectable marker in the generation of transgenic cell lines and organisms.
2
Hygromycin b by Merck Group
Sourced in United States, Germany, United Kingdom, Czechia, Italy, China, France, Sao Tome and Principe
Hygromycin B is a laboratory product manufactured by Merck Group. It is an antibiotic that inhibits protein synthesis in prokaryotic and eukaryotic cells.
3
Fetal bovine serum (fbs) by Thermo Fisher Scientific
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Fetal Bovine Serum (FBS) is a cell culture supplement derived from the blood of bovine fetuses. FBS provides a source of proteins, growth factors, and other components that support the growth and maintenance of various cell types in in vitro cell culture applications.
4
Lipofectamine 2000 by Thermo Fisher Scientific
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Lipofectamine 2000 is a cationic lipid-based transfection reagent designed for efficient and reliable delivery of nucleic acids, such as plasmid DNA and small interfering RNA (siRNA), into a wide range of eukaryotic cell types. It facilitates the formation of complexes between the nucleic acid and the lipid components, which can then be introduced into cells to enable gene expression or gene silencing studies.
5
Hygromycin b by Roche
Sourced in United States, Germany, Switzerland, China
Hygromycin B is a broad-spectrum antibiotic produced by the bacterium Streptomyces hygroscopicus. It functions as a selective agent for eukaryotic cells that have been transfected with the hygromycin B phosphotransferase gene, which confers resistance to the antibiotic.
6
Dulbecco modified eagle medium (dmem) by Thermo Fisher Scientific
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DMEM (Dulbecco's Modified Eagle's Medium) is a cell culture medium formulated to support the growth and maintenance of a variety of cell types, including mammalian cells. It provides essential nutrients, amino acids, vitamins, and other components necessary for cell proliferation and survival in an in vitro environment.
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Penicillin streptomycin by Thermo Fisher Scientific
Sourced in United States, Germany, United Kingdom, China, Canada, France, Japan, Australia, Switzerland, Israel, Italy, Belgium, Austria, Spain, Gabon, Ireland, New Zealand, Sweden, Netherlands, Denmark, Brazil, Macao, India, Singapore, Poland, Argentina, Cameroon, Uruguay, Morocco, Panama, Colombia, Holy See (Vatican City State), Hungary, Norway, Portugal, Mexico, Thailand, Palestine, State of, Finland, Moldova, Republic of, Jamaica, Czechia
Penicillin/streptomycin is a commonly used antibiotic solution for cell culture applications. It contains a combination of penicillin and streptomycin, which are broad-spectrum antibiotics that inhibit the growth of both Gram-positive and Gram-negative bacteria.
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Hygromycin b by InvivoGen
Sourced in United States, France, Hong Kong
Hygromycin B is a broad-spectrum antibiotic used as a selection marker in genetic engineering applications. It inhibits protein synthesis in prokaryotic and eukaryotic cells.
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Streptomycin by Thermo Fisher Scientific
Sourced in United States, United Kingdom, Germany, China, France, Canada, Australia, Japan, Switzerland, Italy, Belgium, Israel, Austria, Spain, Netherlands, Poland, Brazil, Denmark, Argentina, Sweden, New Zealand, Ireland, India, Gabon, Macao, Portugal, Czechia, Singapore, Norway, Thailand, Uruguay, Moldova, Republic of, Finland, Panama
Streptomycin is a broad-spectrum antibiotic used in laboratory settings. It functions as a protein synthesis inhibitor, targeting the 30S subunit of bacterial ribosomes, which plays a crucial role in the translation of genetic information into proteins. Streptomycin is commonly used in microbiological research and applications that require selective inhibition of bacterial growth.
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Puromycin by Merck Group
Sourced in United States, Germany, China, Sao Tome and Principe, United Kingdom, Macao, Canada, Japan, Italy, Switzerland, France, Israel, Australia, Spain, Belgium, Morocco, Sweden
Puromycin is a laboratory product manufactured by Merck Group. It functions as an antibiotic that inhibits protein synthesis in eukaryotic cells.

More about "Hygromycin B"

Hygromycin B is a widely used antibiotic in molecular biology and cell culture applications.
Produced by the bacterium Streptomyces hygroscopicus, this selective marker is commonly utilized to identify and isolate cells that have been successfully transfected or transformed.
Hygromycin B works by inhibiting protein synthesis in eukaryotic cells, making it an effective tool for research.
It is often used in conjunction with other reagents like Fetal Bovine Serum (FBS), Lipofectamine 2000, Dulbecco's Modified Eagle Medium (DMEM), Penicillin/Streptomycin, and Puromycin to optimize cell culture and genetic engineering experiments.
Researchers can leverage the power of AI-powered tools like PubCompare.ai to enhance their Hygromycin B research.
These platforms can help identify optimized protocols from literature, preprints, and patents, ensuring reproducibility and accuracy in their work.
By incorporating Hygromycin B into their research workflows, scientists can effectively select for and isolate successfully modified cells, advancing their studies in molecular biology and beyond.