Nucleic Acids Research, 1984, Vol. 12, No. 14 5707-5717
© 1984
Articles |
High level transient expression of a chloramphenicol acetyl transferase gene by DEAE-dextran mediated DNA transfection coupled with a dimethyl sulfoxide or glycerol shock treatment
Department of Biological Chemistry, Johns Hopkins University School of Medicine , 725 North Wolfe Street, Baltimore, MD 21205, USA
Received April 2, 1984. Revised June 18, 1984. Accepted June 18, 1984.
Using a plasmid containing the bacterial chloramphenicol acetyl transferase gene, we have assayed for transient expression of DNA introduced into mouse L cells by a variety of transfection conditions. High efficiency uptake and expression of this foreign DNA have been achieved by modifying the DEAE dextran mediated trsnafection procedure of McCutchan and Pagano (1) to include a shock with either dimethyl sulfoxide or glycerol. Inclusion of the shock step can increase expreasion of the transfected gene a surprising {small tilde}50 fold. With plasmid constructs that do not replicate after transfection, we can readily detect CAT activity in an overnight autoradiographic exposure from less than 0.1% of an extract from a 60 mm dish of transfected cells. We have determined the amounts of DNA, the amount and time course of DEAE-dextran and dimethyl aulfoxide treatments, the effects of additional DNA, and the time after transfection which yield maximal expression. Overall, this transfection protocol using DEAE-dextrsn coupled to a shock treatment is simple, straightforward, and gives consistently high levels of expression of the input DNA.
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V. Krishnan, G. Elberg, M.-J. Tsai, and S. Y. Tsai Identification of a Novel Sonic Hedgehog Response Element in the Chicken Ovalbumin Upstream Promoter-Transcription Factor II Promoter Mol. Endocrinol., September 1, 1997; 11(10): 1458 - 1466. [Abstract] [Full Text]
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W. Gong, S. Chávez, and M. Beato Point Mutation in the Ligand-Binding Domain of the Progesterone Receptor Generates a Transdominant Negative Phenotype Mol. Endocrinol., September 1, 1997; 11(10): 1476 - 1485. [Abstract] [Full Text]
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C. E. Hughes, F. H. Buttner, B. Eidenmuller, B. Caterson, and E. Bartnik Utilization of a Recombinant Substrate rAgg1 to Study the Biochemical Properties of Aggrecanase in Cell Culture Systems J. Biol. Chem., August 8, 1997; 272(32): 20269 - 20274. [Abstract] [Full Text] [PDF]
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R. Jockers, L. Petit, I. Lacroix, P. d. Coppet, P. Barrett, P. J. Morgan, B. Guardiola, P. Delagrange, S. Marullo, and A. D. Strosberg Novel Isoforms of Mel1c Melatonin Receptors Modulating Intracellular Cyclic Guanosine 3',5'-Monophosphate Levels Mol. Endocrinol., July 1, 1997; 11(8): 1070 - 1081. [Abstract] [Full Text]
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L. Cupillard, K. Koumanov, M.-G. Mattei, M. Lazdunski, and G. Lambeau Cloning, Chromosomal Mapping, and Expression of a Novel Human Secretory Phospholipase A2 J. Biol. Chem., June 20, 1997; 272(25): 15745 - 15752. [Abstract] [Full Text] [PDF]
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K. K. Kuropatwinski, C. De Imus, D. Gearing, H. Baumann, and B. Mosley Influence of Subunit Combinations on Signaling by Receptors for Oncostatin M, Leukemia Inhibitory Factor, and Interleukin-6 J. Biol. Chem., June 13, 1997; 272(24): 15135 - 15144. [Abstract] [Full Text] [PDF]
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H. Kim and H. Baumann The Carboxyl-terminal Region of STAT3 Controls Gene Induction by the Mouse Haptoglobin Promoter J. Biol. Chem., June 6, 1997; 272(23): 14571 - 14579. [Abstract] [Full Text] [PDF]
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T. Zarkowska and S. Mittnacht Differential Phosphorylation of the Retinoblastoma Protein by G1/S Cyclin-dependent Kinases J. Biol. Chem., May 9, 1997; 272(19): 12738 - 12746. [Abstract] [Full Text] [PDF]
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M. Salinas, J. de Weille, E. Guillemare, M. Lazdunski, and J.-P. Hugnot Modes of Regulation of Shab K+ Channel Activity by the Kv8.1 Subunit J. Biol. Chem., March 28, 1997; 272(13): 8774 - 8780. [Abstract] [Full Text] [PDF]
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D. W. White, K. K. Kuropatwinski, R. Devos, H. Baumann, and L. A. Tartaglia Leptin Receptor (OB-R) Signaling. CYTOPLASMIC DOMAIN MUTATIONAL ANALYSIS AND EVIDENCE FOR RECEPTOR HOMO-OLIGOMERIZATION J. Biol. Chem., February 14, 1997; 272(7): 4065 - 4071. [Abstract] [Full Text] [PDF]
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R. V. Rao, B. F. Roettger, E. M. Hadac, and L. J. Miller Roles of Cholecystokinin Receptor Phosphorylation in Agonist-Stimulated Desensitization of Pancreatic Acinar Cells and Receptor-Bearing Chinese Hamster Ovary Cholecystokinin Receptor Cells Mol. Pharmacol., February 1, 1997; 51(2): 185 - 192. [Abstract] [Full Text] [PDF]
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Z. Huang, Y. Chen, S. Pratt, T.-H. Chen, T. Bambino, R. A. Nissenson, and D. M. Shoback The N-terminal Region of the Third Intracellular Loop of the Parathyroid Hormone (PTH)/PTH-related Peptide Receptor Is Critical for Coupling to cAMP and Inositol Phosphate/Ca2+ Signal Transduction Pathways J. Biol. Chem., December 27, 1996; 271(52): 33382 - 33389. [Abstract] [Full Text] [PDF]
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S. Tamai, A. Waheed, L. B. Cody, and W. S. Sly Gly-63 right-arrow Gln substitution adjacent to His-64 in rodent carbonic anhydrase IVs largely explains their reduced activity PNAS, November 26, 1996; 93(24): 13647 - 13652. [Abstract] [Full Text] [PDF]
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S. Kuroda, M. Fukata, K. Kobayashi, M. Nakafuku, N. Nomura, A. Iwamatsu, and K. Kaibuchi Identification of IQGAP as a Putative Target for the Small GTPases, Cdc42 and Rac1 J. Biol. Chem., September 20, 1996; 271(38): 23363 - 23367. [Abstract] [Full Text] [PDF]
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F. P. Limbourg, H. Stadtler, G. Chinnadurai, P. A. Baeuerle, and M. L. Schmitz A Hydrophobic Region within the Adenovirus E1B 19kDa Protein Is Necessary for the Transient Inhibition of NF-kappa B Activated by Different Stimuli J. Biol. Chem., August 23, 1996; 271(34): 20392 - 20398. [Abstract] [Full Text] [PDF]
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L.-H. Wang, N. Matijevic-Aleksic, P.-Y. Hsu, K.-H. Ruan, K. K. Wu, and R. J. Kulmacz Identification of Thromboxane A2 Synthase Active Site Residues by Molecular Modeling-guided Site-directed Mutagenesis J. Biol. Chem., August 16, 1996; 271(33): 19970 - 19975. [Abstract] [Full Text] [PDF]
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J. M. Shields, R. J. Christy, and V. W. Yang Identification and Characterization of a Gene Encoding a Gut-enriched Kruppel-like Factor Expressed during Growth Arrest J. Biol. Chem., August 16, 1996; 271(33): 20009 - 20017. [Abstract] [Full Text] [PDF]
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M. H. Holtmann, S. Ganguli, E. M. Hadac, V. Dolu, and L. J. Miller Multiple Extracellular Loop Domains Contribute Critical Determinants for Agonist Binding and Activation of the Secretin Receptor J. Biol. Chem., June 21, 1996; 271(25): 14944 - 14949. [Abstract] [Full Text] [PDF]
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C.-F. Lai, J. Ripperger, K. K. Morella, J. Jurlander, T. S. Hawley, W. E. Carson, T. Kordula, M. A. Caligiuri, R. G. Hawley, G. H. Fey, et al. Receptors for Interleukin (IL)-10 and IL-6-type Cytokines Use Similar Signaling Mechanisms for Inducing Transcription through IL-6 Response Elements J. Biol. Chem., June 14, 1996; 271(24): 13968 - 13975. [Abstract] [Full Text] [PDF]
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T.-F. Wang and G. Guidotti CD39 Is an Ecto-(Ca[IMAGE],Mg[IMAGE])-apyrase J. Biol. Chem., April 26, 1996; 271(17): 9898 - 9901. [Abstract] [Full Text] [PDF]
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P. R. Turner, T. Bambino, and R. A. Nissenson A Putative Selectivity Filter in the G-protein-coupled Receptors for Parathyroid Hormone and Secretin J. Biol. Chem., April 19, 1996; 271(16): 9205 - 9208. [Abstract] [Full Text] [PDF]
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T. Kordula, J. Ripperger, K. K. Morella, J. Travis, and H. Baumann Two Separate Signal Transducer and Activator of Transcription Proteins Regulate Transcription of the Serine Proteinase Inhibitor-3 Gene in Hepatic Cells J. Biol. Chem., March 22, 1996; 271(12): 6752 - 6757. [Abstract] [Full Text] [PDF]
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J. C. Otto and P. J. Casey The Hepatitis Delta Virus Large Antigen Is Farnesylated Both in Vitro and in Animal Cells J. Biol. Chem., March 1, 1996; 271(9): 4569 - 4572. [Abstract] [Full Text] [PDF]
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F. Ozcelebi, R. V. Rao, E. Holicky, B. J. Madden, D. J. McCormick, and L. J. Miller Phosphorylation of Cholecystokinin Receptors Expressed on Chinese Hamster Ovary Cells J. Biol. Chem., February 16, 1996; 271(7): 3750 - 3755. [Abstract] [Full Text] [PDF]
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E. Zvaritch, Gér. Lambeau, and M. Lazdunski Endocytic Properties of the M-type 180-kDa Receptor for Secretory Phospholipases A(2) J. Biol. Chem., January 5, 1996; 271(1): 250 - 257. [Abstract] [Full Text] [PDF]
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R. G. Boot, G. H. Renkema, A. Strijland, A. J. van Zonneveld, and J. M. F. G. Aerts Cloning of a cDNA Encoding Chitotriosidase, a Human Chitinase Produced by Macrophages J. Biol. Chem., November 3, 1995; 270(44): 26252 - 26256. [Abstract] [Full Text] [PDF]
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C.-F. Lai, J. Ripperger, K. K. Morella, Y. Wang, D. P. Gearing, N. D. Horseman, S. P. Campos, G. H. Fey, and H. Baumann STAT3 and STAT5B Are Targets of Two Different Signal Pathways Activated by Hematopoietin Receptors and Control Transcription via Separate Cytokine Response Elements J. Biol. Chem., October 6, 1995; 270(40): 23254 - 23257. [Abstract] [Full Text] [PDF]
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M. L. Schmitz, M. A. d. S. Silva, and P. A. Baeuerle Transactivation Domain 2 (TA[IMAGE]) of p65 NF-[IMAGE]B J. Biol. Chem., June 30, 1995; 270(26): 15576 - 15584. [Abstract] [Full Text] [PDF]
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C.-F. Lai, J. Ripperger, K. K. Morella, Y. Wang, D. P. Gearing, G. H. Fey, and H. Baumann Separate Signaling Mechanisms Are Involved in the Control of STAT Protein Activation and Gene Regulation via the Interleukin 6 Response Element by the Box 3 Motif of gp130 J. Biol. Chem., June 23, 1995; 270(25): 14847 - 14850. [Abstract] [Full Text] [PDF]
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M. H. Holtmann, E. M. Hadac, and L. J. Miller Critical Contributions of Amino-terminal Extracellular Domains in Agonist Binding and Activation of Secretin and Vasoactive Intestinal Polypeptide Receptors J. Biol. Chem., June 16, 1995; 270(24): 14394 - 14398. [Abstract] [Full Text] [PDF]
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J. Halstead, K. Kemp, and R. A. Ignotz Evidence for Involvement of Phosphatidylcholine-Phospholipase C and Protein Kinase C in Transforming Growth Factor-[IMAGE] Signaling J. Biol. Chem., June 9, 1995; 270(23): 13600 - 13603. [Abstract] [Full Text] [PDF]
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H. Weng, S.-Y. Choi, and D. V. Faller The Moloney Leukemia Retroviral Long Terminal Repeat trans-Activates AP-1-inducible Genes and AP-1 Transcription Factor Binding J. Biol. Chem., June 9, 1995; 270(23): 13637 - 13644. [Abstract] [Full Text] [PDF]
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A. Iida-Klein, J. Guo, L. Y. Xie, H. Jüppner, J. T. Potts Jr., H. M. Kronenberg, F. R. Bringhurst, A. B. Abou-Samra, and G. V. Segre Truncation of the Carboxyl-terminal Region of the Rat Parathyroid Hormone (PTH)/PTH-related Peptide Receptor Enhances PTH Stimulation of Adenylyl Cyclase but Not Phospholipase C J. Biol. Chem., April 14, 1995; 270(15): 8458 - 8465. [Abstract] [Full Text] [PDF]
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K. K. Morella, C.-f. Lai, S. Kumaki, N. Kumaki, Y. Wang, E. M. Bluman, B. A. Witthuhn, J. N. Ihle, J. Giri, D. P. Gearing, et al. The Action of Interleukin-2 Receptor Subunits Defines a New Type of Signaling Mechanism for Hematopoietin Receptors in Hepatic Cells and Fibroblasts J. Biol. Chem., April 7, 1995; 270(14): 8298 - 8310. [Abstract] [Full Text] [PDF]
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