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J. Biol. Chem. 284, 6773–6781. Acute lymphoblastic leukemia-associated JAK1 mutants activate the Janus kinase/STAT pathway via interleukin-9 receptor alpha homodimers. 2009

Hornakova, T., Staerk, J., Royer, Y., Flex, E., Tartaglia, M., Constantinescu, S.N., Knoops, L. and Renauld, J.C.

Notes: The authors studied the ability of JAK1 V658F and A634D mutants to activate the Janus kinase (JAK)/STAT pathway when expressed alone or together with the other components of the interleukin-9 receptor complex. The BOX1 motif of wild-type IL-9Rα, the JAK interacting region, was mutated from PXP to SXS using the GeneEditor™ in vitro Site-Directed Mutagenesis System. To assess STAT transcriptional activity, HEK293 human embryonic kidney, COS-7 monkey kidney, U4C human fibrosarcoma and g2A cells were cotransfected with 250ng of the appropriate constructs, 500ng of firefly luciferase vectors and 50ng of pRL-TK Vector and empty plasmid for a total 1.5µg of DNA. After 24 hours, the cells were lysed in 150µl of Passive Lysis Buffer and reporter activity measured using the Dual-Luciferase® Reporter Assay System. The ProFection® Mammalian Transfection System—Calcium Phosphate was used to transfect 106 HEK293 cells in a six-well plate with 3.75µg of plasmid for Western blot analysis and cotransfected 6 × 106 HEK293 cells in a 100mm dish with 14µg plasmid for immunoprecipitation studies. (4025)

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J. Immunol. 172, 2687-2696. HIV-1 does not provoke alteration of cytokine gene expression in lymphoid tissue after acute infection ex vivo. 2004

Audige, A., Schlaepfer, E., Bonanomi, A., Joller, H., Knuchel, M.C., Weber, M., Nadal, D. and Speck, R.F.

Notes: The authors used real-time quantitative PCR to characterize cytokine response after HIV infection of human lymphoid tissues. To synthesize first-strand cDNA, total RNA was reverse transcribed using the ImProm-II™ Reverse Transcription System: 200U of ImProm-II Reverse Transcriptase, 2µg of total RNA, 500ng of oligo(dT) primer, 500µM dNTPs 3mM MgCl2 and 24U of RNase inhibitor in 1x ImProm-II™ reaction buffer. To obtain viral stocks for infection, 293T cells were transfected with the proviral plasmids pNL4-3 and pYU-2 using the ProFection® Mammalian Transfection System—Calcium Phosphate. (3455)

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J. Biol. Chem. 276, 31786-31792. The Drosophila U1 and U6 gene proximal sequence elements act as important determinants of the RNA polymerase specificity of small nuclear RNA gene promoter in vitro and in vivo. 2001

McNamara-Schroeder, K.J., Hennessey, R.F., Harding, G.A., Jensen, R.C. and Stumph, W.E.

Notes: Drosophila S2 cells were transfected with a U6 maxigene plasmid and a control firefly luciferase construct (derived from the pGL2 Basic Vector) using the ProFection® Mammalian Transfection System-Calcium Phosphate. Expression of the maxigene was confirmed by isolating total RNA from an aliquot of the cells using the RNAgents® Total RNA Isolation System and performing primer extension analysis with the aid of the Primer Extension System. Luciferase assays were performed on an aliquot of cells after lysis with Reporter Lysis Buffer. (2576)

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J. Cell Biol. 150, 165-175. Glutamate slows axonal transport of neurofilaments in transfected neurons. 2000

Ackerley, S., Grierson, A.J., Brownlees, J., Thornhill, P., Anderton, B.H., Leigh, P.N., Shaw, C.E., and Miller C.C.

Notes: The authors seek to determine the role of glutamate in excitotoxicity and neurofilament accumulation seen in some neurodegenerative diseases. Neurofilament light, middle, and heavy chains were expressed from rat cDNAs cloned into the pCI-neo Mammalian Expression vector in SW-13 cells. Primary rat cortical neurons were transfected with a neurofilament middle chain and green fluorescent fusion protein. SW13 cells and primary rat cortical neurons were transfected with the ProFection® Mammalian Transfection System–Calcium Phosphate. The CytoTox 96® Non-Radioactive Cytotoxicity Assay was used to monitor glutamate toxicity in these cell. To determine the role of the MAPK and JNK signaling pathways,  SW13- cells and primary neuronal cells were immunostained for dually phosphorylated MAPK and JNK using Promega's Anti-ACTIVE® MAPK pAb and Anti-ACTIVE® JNK pAb, respectively. Cells were fixed in 4% paraformaldehyde, permeabilized in 0.1% Triton® X-100 in PBS, blocked with  0.2% Tween® 20 in TBS, and incubated with primary antibodies diluted in blocking solution. Western blot analyses were performed on the primary cortical neurons to quantitate the level of dually phosphorylated MAPK protein  The blots were also probed with a pan MAPK antibody that detects total (active and inactive) MAPK. (2382)

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Gene Ther. 5, 65-75. Inhibition of HIV-1 replication by combined expression of gag dominant negative mutant and a human ribonuclease in a tightly controlled HIV-1 inducible vector 1998

Cara, A., Rybak, S.M., Newton, D.L., Rottschafer, S.E., Reitz Jr., M.S., and Gusella, G.L.

Notes: The ProFection® Mammalian Transfection System - Calcium Phosphate was used to transfect HeLa and HeLa-Tat cell lines. The authors developed a vector-antiviral gene system for use in gene therapy in the treatment of HIV-1. pGEM®-luc was used as a negative control in luciferase assays of transfected cells. (1388)

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Hepatology 28, 1147-1153. Mutations in the interferon-sensitivity determining region of hepatitis C virus and transcriptional activity of the nonstructuraql region 5A protein. 1998

Fukuma, T., Enomoto, N., Marumo, F., Sato, C.

Notes: The ProFection® Mammalian Transfection System-Calcium Phosphate was used to transfect reporter plasmids into HuH-7, a human hepatoma cell line. The cells were plated at 1.2 X 106 in a 100mm dish and transfected 48 hours later. Transfection efficiency was monitored with a co-transfection of the pSV-β-Galactosidase Control Vector with the CAT reporter vectors. (1130)

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J. Neurosci. 18, 751-762. Phosphorylation of c-Jun is necessary for apoptosis induced by survival signal withdrawal in cerebellar granule neurons. 1998

Watson, A. , Eilers, A. , Lallemand, D. , Kyriakis, J. , Rubin, L. L. , Ham, J.

Notes: ProFection® Mammalian Transfection System (Calcium Phosphate) was used to transfect primary cerebellar granule neurons from 8 day old Sprague Dawley rats. The complete details of the transfection is provided, including the efficiency of the transfection. Efficiencies, as determined by transfected control β-galactosidase, ranged from 3-7%. (0170)

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Hepatology 28, 1013-1021. The hepatitis B virus X protein up-regulates tumor necrosis factor alpha gene expression in hepatocytes. 1998

Lara-Pezzi, E., Majano, P.L., Gómez-Gonzalo, M., García-Monzón, C., Moreno-Otero, R., Levrero, M., López-Cabrera, M.

Notes: Transfections of HepG2 cells, 2.2.15 (a Hepatitis B-infected cell line) and HeLa cells were accomplished with the ProFection® Mammalian Transfection System-Calcium Phosphate. Transfection efficiencies were monitored with co-transfected pSV-β-Galactosidase Control Vector, but the efficiencies were not reported. (0843)

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J. Biol. Chem. 272, 26918-26925. Detection and characterization of Sp1 binding activity in human chondrocytes and its alterations during chondrocyte dedifferentiation 1997

Dharmavaram, R.M. , Liu, G. , Mowers, S.D. , Jimenez, S.A.

Notes: The Profection® Mammalian Transfection System (CaPO4) was used for transient transfection of the Drosophila Schneider line 2 cells. A lot of detail is provided. (1230)

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Blood 89, 1394-1404. Detection of anaplastic lymphoma kinase (ALK) and nucleolar protein nucleophosmin (NPM)-ALK proteins in normal and neoplastic cells with the monoclonal antibody ALK1. 1997

Pulford, K., Lamant, L., Morris, S.W., Butler, L.H., Wood, K.M., Stroud, D., Delsol, G., Mason, D.Y.

Notes: The ProFection® Mammalian Transfection System-CaPO4 was used to transiently transfect 293T cells. (0521)

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J. Cell Biol. 136(5), 1059-1070. Isoforms of ankyrin-3 that lack the NH2-terminal repeats associate with mouse macrophage lysosomes. 1997

Hoock, T.C., Peters, L.L. and Lux, S.E.

Notes: The inhibitor was used to protect RNA during RT-PCR. (2246)

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J. Biol. Chem. 272, 20251-20258. Modulation of interleukin (IL)-13 binding and signaling by the gamma chain of the IL-2 Receptor 1997

Obiri, N. I. , Murata, T. , Debinski, W. , Puri, R. K.

Notes: The ProFection® Mammalian Transfection System-CaPO4 transfection system was used to generate stably transfected ML-RCC renal carcinoma cells. (0628)

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Proc. Natl. Acad. Sci. USA 94, 11567-11572. Phenotypic knockout of HIV type 1 chemokine coreceptor CCR-5 by intrakines as potential therapeutic approach for HIV-1 infection 1997

Yang, A. G. , Bai, X. , Huang, X. F. , Yao, C. , Chen, S.

Notes: The ProFection® Mammalian Transfection System-Calcium Phosphate was used to transiently transfect HeLa-T4+ cells at 50% confluency. (0138)

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J. Biol. Chem. 271, 22746-22753. Identification of a novel cis-acting negative regulatory element affecting expression of the CYP1A1 gene in rat epidermal cells. 1996

Walsh, A. A., Tullis, K., Rice, R. H., Denison, M. S.

Notes: Expression of luciferase reporter constructs, derived from the pGL2-Basic Vector, were studied in rat epidermal keratinocytes. The ProFection® Mammalian Transfection System (calcium phosphate) was used as well. (0194)

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