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J. Gen. Virol. 93, 2408–18. Evolution of the hepatitis E virus hypervariable region. 2012

Smith, D.B., Vanek, J., Ramalingam, S., Johannessen, I., Templeton, K. and Simmonds, P.

Notes: The authors of this study investigated the function of the hypervariable region (HVR) present in open reading frame 1 (ORF1) in the hepatitis E virus (HEV) by measuring the diversity of the HVR in HEV samples from acutely infected patients and in epidemiologically related samples. They sequenced HEV HVR PCR products from limited dilution cDNA from 8 patients PCR positive for ORF2 of HEV. HEV RNA was extracted from serum using a commercial kit, and then HEV RNA was amplified using the Access RT-PCR System. A second round of PCR was performed using GoTaq® polymerase. cDNA was generated using random hexamer or appropriate primers in the presence of Recombinant RNasin® Ribonuclease Inhibitor. (4242)

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mBio. 3(5), e00266–12. A multicenter blinded analysis indicates no association between chronic fatigue syndrome/myalgic encephalomyelitis and either xenotropic murine leukemia virus-related virus or polytropic murine leukemia virus. 2012

Alter, H.J., Mikovits, J.A., Switzer, W.M., Ruscetti, F.W., Lo, S.C., Klimas, N., Komaroff, A.L., Montoya, J.G., Bateman, L., Levine, S., Peterson, D., Levin, B., Hanson, M.R., Genfi, A., Bhat, M., Zheng, H., Wang, R., Li, B., Hung, G.C., Lee, L.L., Sameroff, S., Heneine, W., Coffin, J., Hornig, M. and Lipkin, W.I.

Notes: In this report, the original investigators who found XMRV and pMLV (polytropic murine leukemia virus) in blood of subjects with Chronic Fatigue Syndrome (CFS) report that this association is not confirmed in a blinded analysis of samples from rigorously characterized subjects.

The CDC performed nucleic acid testing assays. Plasma was centrifuged and RNA isolated from the pellet. Quantitative real-time RT-PCR assays (qRT-PCR) for generic pMLV/XMRV pro (protease) and gag detection were performed on RNA extracts, using the AccessQuick™ RT-PCR System and an AgPath one-step RT-PCR kit.

ArrayScript RT and AmpliTaq Gold DNA polymerase were used for cDNA synthesis and amplification in the pro and gag qRT-PCR assays, respectively. A third PCR was done using the primers XPOLOF and XPOLOR, followed by a nested PCR with the primers XPOLIF and XPOLIR for the generic detection of MLV/XMRV 216-bp pol sequences. For this reaction, cDNA synthesis and amplification of RNA was done using Promega AMV Reverse Transcriptase and a RobustI RT-PCR kit. Each PCR experiment included 20 water-only reactions to control for contamination. (4300)

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Nucl. Acids Res. 38, e167. In situ reverse transcription: the magic of strength and anonymity. 2010

Ligasová, A., and Koberna, K.

Notes: This paper describes a method for detection of polyA RNA in permeablized cells and cell sections. The method is based on incorporation of 5-bromo-2´-deoxyuridine (BrdUTP) into cDNA by AMV reverse transcriptase. The BrdUTP was easily detectable in DNA-RNA duplexes, and undetectable in DNA-DNA duplexes, making it possible to use the method to detect RNA in situ. RNasin® Ribonuclease Inhibitor was used in the reverse transcriptase reaction mix. (4226)

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Am. J. Pathol. 171, 1153-1167. Dissecting the impact of chemotherapy on the human hair follicle: a pragmatic in vitro assay for studying the pathogenesis and potential management of hair follicle dystrophy. 2007

Bodó, E., Tobin, D.J., Kamenisch, Y., Bíró, T., Berneburg, M., Funk, W. and Paus, R.

Notes: To study how chemotherapy affects hair follicles (HF), the researchers microdissected and cultured human anagen VI scalp follicles and treated the cells with 4-Hydroperoxycyclophosphamide (4-HC) at similar concentrations to those received during therapy. After incubation for 24 hours with 30µmol/L of 4-HC, the HFs were homogenized and genomic DNA was purified using the Wizard® SV Genomic DNA Purification System. To determine if the treatment induced the mitochondrial common DNA deletion, the isolated DNA was subjected to real-time PCR. Further examination of gene expression was performed by isolating total RNA from two HF samples, reverse transcribing 3µg of the RNA using 15U of AMV Reverse Transcriptase and 0.025 µg/µl random primers, and amplifying seven human genes in a TaqMan® Assay. (3746)

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Cell Death Differ. 9, 1334-1342. Differential subcellular localization of functionally divergent survivin splice mutants 2002

Mahotka, C., Liebmann, J., Wenzel, M., Suschek, C.V., Schmitt, M., Gabbert, H.E., Gerharz, C.D.

Notes: Promega reverse transcription buffer, AMV reverse transcriptase, and RNAsin® RNAse inhibitor were used in reverse transcription reactions of total RNA from HepG2 heptatoma cells. PCR products were cloned into the pGEMT vector. (2613)

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J. Bacteriol. 184, 4148-4160. DNA-binding activities of the HilC and HilD virulence regulatory proteins of Salmonella enterica serovar Typhimurium. 2002

Olekhnovich, I.N. and Kadner, R.J.

Notes: The RNAgents® Total RNA Isolation System was used to isolate total RNA from 100ml cultures of S. enterica grown to an OD600 of 1.0 in permissive conditions of limited oxygen and high osmolarity. The RNA was used in primer extension analysis with AMV Reverse Transcriptase. (2574)

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J. Biol. Chem. 277, 17349-17358. Structural Determinants of BRCA1 Translational Regulation 2002

Sobczak, K. and Krzyzosiak, W.J.

Notes: The authors investigated expression of BRCA1 from alternative transcripts possessing different 5' UTRs. RT-PCR using Promega's AMV reverse transcriptase was performed to generate cDNAs from total RNA isolated from human tissue. Promega's Taq DNA Polymerase was used for the PCR reaction. The authors also prepared two mRNAs that contained one or the other of the 2 alternative 5' UTRs (ex1a or ex1b) fused with the luciferase coding region. To generate the corresponding DNA for these constructs, the luciferase coding region was amplified from Promega's pGEM vector and ligated to PCR amplified ex1a or exlb. These ligation products served as the template amplifying two cDNA constructs (exla-luc or ex1b-luc).Ten additional cDNAs were made containing a variety of changes to the 5'UTR region of BRCA1. In vitro translation experiments were performed to determine how the composition of the 5'UTR affected protein expression levels (using Promega's RNasin to protect transcribed mRNA; T7 polymerase buffer; and rabbit reticulocyte and wheat germ extracts for translation). The authors conclude that secondary structures associated with elements in the longer 5'UTR reduced translation rates and could be responsible for the reduced expression of BRCA1 often associated with spontaneous ovarian and breast cancers. (2441)

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Antimicrob. Agents Chemother. 45, 1151–1161. Novel Carbapenem-Hydrolyzing β-Lactamase, KPC-1, from a Carbapenem-Resistant Strain of Klebsiella pneumoniae. 2001

Yigit, H., Queenan, A.M., Anderson, G.J., Domenech-Sanchez, A., Biddle, J.W., Steward, C.D., Alberti, S., Bush, K. and Tenover, F.C.

Notes: The study of a carbapenem resistant strain of Klebsiella pneumoniae lead to the description of a novel form of β-lactamase, which confers resistance to imipenem, meropenem, extended-spectrum cepahlosporins, and aztreonam. Total RNA used in a primer extension assay was isolated with the SV Total RNA Isolation System . Primer extension reactions were performed with Promega's Primer Extension System- AMV Reverse Transcriptase to determine the transcriptional start site of the KPC-1 β-lactamase gene. The KPC-1 β-lactamase gene was then sequenced with the fmol® DNA Sequencing System (2300)

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Appl. Environ. Microbiol. 66, 3778–3783. A Serine Protease-Encoding Gene (aprII) of Altermonas sp. Strain O-7 Is Regulated by the Iron Uptake Regulator (Fur) Protein. 2000

Tsujibo, H., Miyamoto, K., Okamoto, T., Orikoshi, H., and Inamori, Y.

Notes: Total RNA was isolated from Alteromonas sp. strain O-7 for Northern blot analysis of the aprII gene to show that aprII is upregulated under iron rich growth conditions. The transcriptional start site of this gene was mapped in a primer extension reaction using Promega's AMV reverse transcriptase. (2304)

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FASEB J. 14, 740–751. Cellular dedifferentiation of endothelium is linked to activation and silencing of certain nuclear transcription factors: Implications for endothelial dysfunction and vascular biology. 2000

Thum, T., Haverich, A. and Borlak, J.

Notes: The SV Total RNA Isolation System was used to isolate total RNA from rat endothelial cells. The isolated RNA was used for RT-PCR and semi-quantitative RT-PCR with cDNA generated with AMV Reverse Transcriptase. (2175)

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Cell 101, 249-258. Functional and genomic analyses reveal an essential coordination between the unfolded protein response and ER-associated degradation. 2000

Travers K.J., Patil C.K., Wodicka L., Lockhart D.J., Weissman J.S. and Walter P.

Notes: Poly A+ RNA was isolated from the total RNA of three different strains of Saccharomyces cerevisiae, wild-type (JC103), CS165 and JC408, using the PolyATtract® mRNA Isolation System. The isolated polyA+ RNA was used to make biotinylated cRNAs via a referenced method using AMV Reverse Transcriptase. The resulting probes were used in microarray analysis. (2696)

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Eur. J. Biochem. 267, 7224–7229. Identification and characterization of a novel transcriptional regulator, MatR, for malonate metabolism in Rhizobium leguminosarum bv. trifolii 2000

Lee, H.Y., An, J.H. and Kim, Y.S.

Notes: Total RNA was isolated from Rhizobium leguminosarum bv. trifolii using the SV Total RNA Isolation System. This RNA was used in a primer extension assay using Promega's AMV Reverse Transcriptase to determine the transcriptional start site of the mat operon. Nested deletions during the sequencing of the mat promoter were prepared using the Erase-a-Base® System. The firefly luciferase gene from pGL3 Basic was used to construct a reporter construct containing the mat promoter to study transcriptional effects of MatR. The interaction of the MatR protein with the mat operator was mapped using the Core Footprinting System. (2308)

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J. Biol. Chem. 274, 21830–21839. Purification, cDNA cloning, and expression of GDP-L-Fuc:Asn-linked GlcNAc alpha1,3-fucosyltransferase from mung beans. 1999

Leiter, H., Mucha, J., Staudacher, E., Grimm, R., Glössl, J. and Altmann, F.

Notes: Total RNA was isolated from 3-day-old mung bean hypocotyls using the SV Total RNA Isolation System. RT for RT-PCR was performed with the Reverse Transcription System. (0825)

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J. Biol. Chem. 273, 27420-27429. Characterization of the muring fatty acid transport protein gene and its insulin response sequence. 1998

Hui, T.Y., Frohnert, B.I., Smith, A.J., Schaffer, J.E., Bernlohr, D.A.

Notes: Luciferase reporter constructs were cotransfected into 3T3-L1 preadipocytes with the pRL-CMV Vector at a 25:1 ratio and luciferase activities were determined with the Dual-Luciferase® Reporter Assay System. RT-PCR was accomplished with AMV Reverse Transcriptase and Taq DNA Polymerase. RNase Protection Assay were performed with the aid of the RNase ONE™ Ribonuclease. (0988)

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Genetics 148, 815-825. Conserved regions of the timeless (tim) clock gene in Drosophila analyzed through phylogenetic and functional studies. 1998

Ousley, A., Zafarullah, K., Chen, Y., Emerson, M., Hickman, L., Sehgal, A.

Notes: For the RT-PCR experiment, total RNA isolated from fly heads was reverse transcribed by Promega AMV-RT and then the cDNA was amplified by Promega Taq DNA polymerase. (0574)

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Am. J. Hum. Genet. 62, 599-609. Identification of novel mutations in the ryanodine-receptor gene (RYR1) in malignant hyperthermia: genotype-phenotype correlation. 1998

Manning, B. M. , Quane, K. A. , Ording, H. , Urwyler, A. , Tegazzin, V. , Lehane, M. , O'Halloran, J. , Hartung, E. , Giblin, L. M. , Lynch, P. J. , Vaughan, P. , Censier, K. , Bendixen, D. , Comi, G. , Heytens, L. , Monsieurs, K. , Fagerlund, T. , Wolz, W. , Heffron, J. J. , Muller, C. R. , McCarthy, T. V.

Notes: First-strand cDNA was synthesized by use of the Reverse Transcription System and random primers. (0728)

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J. Biol. Chem. 273, 27474-27483. Sp1 and Sp3 regulate transcriptional activity of the facilitative glucose transporter isoform-3 gene in mammalian neuroblasts and trophoblasts. 1998

Rajakumar, R.A., Thamotharan, S., Menon, R.K., Devaskar, S.U.

Notes: AMV Reverse Transcriptase was used for primer extension analysis from poly(A)+ RNA. The Riboprobe® in vitro Transcription System was used to generate [32P]-antisense RNA probes for RNase protection assays. Promoter studies were performed in N2A murine neuroblasotoma cells. Experimental constructs were assembled in the pGL2-Basic Vector and cotransfected with the pRL-TK Vector at a 10:1 ratio and luciferase activities determined with the Dual-Luciferase® Reporter Assay System. Some promoter studies were performed in Drosophila Schneider cells with luciferase promoter constructs and an RSV-driven beta-galactosidase vector. Beta-Galactosidase activity was determined with the Beta-Galactosidase Enzyme Assay System. (0531)

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Genetics 150, 643-650. Tetrahymena mutants with short telomeres. 1998

Ahmed, S., Sheng, H., Niu, L. and Henderson, E.

Notes: PCR products were excised, eluted, and precipitated prior to sequencing with the fmol® DNA Cycle Sequencing System. Promega's RNasin® Ribonuclease Inhibitor and AMV Reverse Transcriptase were used in a reverse transcription assay. (2065)

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Biochem. J. 332, 773-780. Transcript heterogeneity of the human reduced folate carrier results from the use of multiple promoters and variable splicing of alternative upstream exons. 1998

Zhang, L. , Wong, S. C. , Matherly, L. H.

Notes: The authors used the PolyATtract® mRNA Isolation System to isolate mRNA from total RNA. They used this mRNA-rich fraction for primer extension analysis using Promega's AMV Reverse Transcriptase. The Wizard® Plus Midiprep DNA Purification System was used for various plasmid isolations and the Erase-a-Base®  System was used to generate deletion series. The Dual-Luciferase® Reporter Assay System was used to study promoters cloned into the pGL3-Basic Vector. The pRL-SV40 was used as a transfection control plasmid. (0095)

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J. Biol. Chem. 273, 5607-5614. Transcriptional activation of the UDP-glucuronosyltransferase 1A7 gene in rat liver by aryl hydrocarbon receptor ligands and oltipraz. 1998

Metz, R.P. and Ritter, J.K

Notes: The RNasin Ribonuclease Inhibitor was used to protect RNA transcripts during primer extension analysis and in vitro transcription of RNA probes. The resultant RNA probes were used for RNase protection assays. (1639)

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Microbiology 94, 6484-6489. A cluster of bacterial genes for anaerobic benzene ring biodegradation. 1997

Egland, P.G., Pelletier, D.A., Marilyn, D.M., Gibson, J. and Harwood, C.S

Notes: The authors used Promega's AMV Primer Extension System and Access RT-PCR System for this study. (2216)

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Proc. Natl. Acad. Sci. USA 94, 6216-6221. A new member of the tumor necrosis factor/nerve growth factor receptor family inhibits T cell receptor–induced apoptosis. 1997

Nocentini, G., Giunchi, L., Ronchetti, S., Krausz, L.T., Bartoli, A., Moraca, R., Migliorati, G. and Riccardi, C.

Notes: The TNT® Coupled Transcription/Translation System was used to verify the 25.3kDa size of the cloned receptor. AMV reverse transcriptase was used for first strand cDNA synthesis prior to PCR. (1520)

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Br. J. Haematol. 96(1), 183-185. Efficient RT-PCR on platelet mRNA after long-term storage. 1997

Peyruchaud, O., Nurden, A. and Bourre, F.

Notes: Promega's AMV Reverse Transcriptase and Taq DNA Polymerase were used to perform RT-PCR in this study. (2021)

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J. Biol. Chem. 272, 16364-16373.. Expression of mixed lineage kinase-1 in pancreatic beta-cell lines at different stages of maturation and during embryonic pancreas development 1997

DeAizpurua, H. J. , Cram, D. S. , Naselli, G. , Devereux, L. , Dorow, D. S.

Notes: Poly(A)+ RNA was isolated from RIN rat insulinoma cell total RNA with PolyATtract® mRNA Isolation System III. (1264)

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J. Biol. Chem. 272(26), 16364-16373. Expression of mixed lineage kinase-1 in pancreatic beta-cell lines at different stages of maturation and during embryonic pancreas development. 1997

DeAixpurua, H.J., Cram, D.S., Naselli, G., Devereux, L. and Dorow, D.S.

Notes: Poly(A+) RNA was isolated from RIN rat insulinoma cell total RNA and was used for Northern blots. The authors used Promega's PolyATtract® mRNA Isolation System and AMV Reverse Transcriptase in this study. (2107)

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