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J. Cell Biol. 176, 473–482. Chlamydomonas reinhardtii hydin is a central pair protein required for flagellar motility. 2007

Lechtreck, K.F. and Witman, G.B.

Notes: The authors knocked down expression of the hydin gene, which encodes a flagellar protein, to determine the function and location of hydin in Chlamydomonas reinhardtii. Amplification steps in the creation of the RNAi expression vector to target hydin expression were performed using GoTaq® Flexi DNA Polymerase. (3723)

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Am. J. Pathol. 171, 19–31. A sertoli cell-specific knockout of connexin43 prevents initiation of spermatogenesis. 2007

Brehm, R., Zeiler, M., Rüttinger, C., Herde, K., Kibschull, M., Winterhager, E., Willecke, K., Guillou, F., Lécureuil, C., Steger, K., Konrad, L., Biermann, K., Failing, K. and Bergmann, M.

Notes: To study the role of connexin42 (cx43) in testis development, the authors generated a conditional cx43 knockout mouse, which lacked the cx43 gene in Sertoli cells. To confirm that the cx43 gene was deleted in these mice, PCR was performed using primers specific to cx43, 1X Colorless GoTaq® Flexi Reaction Buffer, 2mM MgCl2, dNTPs and 0.15µl of GoTaq® DNA Polymerase. Tissue-specific deletion of cx43 was confirmed by amplifying RNA isolated from mouse testis, heart and tail was also confirmed using the same PCR components. (3713)

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Proc. Natl. Acad. Sci. USA 104, 10637–10642. Commensal and pathogenic Escherichia coli use a common pilus adherence factor for epithelial cell colonization. 2007

Rendón, M.A., Saldaña, Z., Erdem, A.L., Monteiro-Neto, V., Vázquez, A., Kaper, J.B., Puente, J.L. and Girón, J.A.

Notes: The authors identified an adherence factor of enterohemorrhagic E. coli that is involved in colonization of cultured epithelial cells. This factor, named E. coli common pilus (ECP), is encoded by the ecpA gene, which is present 96% of E. coli strains tested, as determined by PCR. The remaining 4% of the strains were found to be deficient in the ECP operon, as determined by multiplex PCR amplification of ecpR, ecpA, epcB and ecpC sequences. PCR were performed using GoTaq® Green Master Mix. An ecpA deletion mutant exhibited impaired adherence compared to the wildtype E. coli strain. Complementation of the mutant strain with the plasmid pMR13, the pGEM®-T Vector cotaining the ecpA gene, restored the strain's ability to adhere to epithelial cells. (3719)

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J. Biol. Chem. 282, 21798–21809. Control and regulation of KplE1 prophage site-specific recombination: a new recombination module analyzed. 2007

Panis, G., Méjean, V. and Ansaldi, M.

Notes: The authors studied the defective prophage KplE1 in E. coli K12 to map the binding sites of proteins required for recombination. Prior to in vivo excision assays in two E. coli K12 strains, the presence of three DNA sequences required for recombination was confirmed by PCR using GoTaq® DNA Polymerase. In vitro excision assays were also performed using linear and supercoiled DNA substrates that were purified using the Wizard® PCR Clean-Up System. Finally the phage-encoded integraseS (IntS) mRNA was quantitated by real-time RT-PCR. The RNA template was purified from E. coli K12 using the PureYield™ RNA Midiprep System. (3722)

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J. Biol. Chem. 282, 21818–21828. Degradation of hsp70 and other mRNAs in Drosophila via the 5´ 3´ pathway and its regulation by heat shock. 2007

Bönisch, C., Temme, C., Moritz, B. and Wahle, E.

Notes: The authors studied hsp70 mRNA degradation in Drosophila Schneider cells. mRNA deadenylation and decay were monitored by Northern blot. Two of the Northern blot probes used to visualize the mRNA decay products were synthesized by transcription of linearized plasmids using T7 RNA Polymerase and [α-32P] UTP. A population of deadenylated mRNA was created by hybridizing mRNA with oligo(dT) and treating with RNase H. CCR4•NOT was identified as the main deadenylase involved in mRNA decay, and the PAN2:PAN3 deadenylase was a minor contributor. RNA interference was used to knock down expression of PAN2 and CAF1, a subunit of CCR4•NOT, to assess the effect on mRNA decay. Reduced expression levels of PAN2 and CAF1 were confirmed by semi-quantitative RT-PCR and Western blotting, respectively. RT-PCR was performed using 1.5µg total RNA and 150 units of MMLV Reverse Transcriptase in a 25µl reaction. One microliter of the RT reaction was used as a template in an 80µl PCR using 0.5 units of GoTaq® DNA Polymerase, 1.5mM MgCl2 and 1X Green GoTaq® Flexi Reaction Buffer. (3707)

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J. Biol. Chem. 282, 29847–29854. Differential regulation of vitamin D receptor (VDR) by the p53 family: p73-dependent induction of VDR upon DNA damage. 2007

Kommagani, R., Payal, V. and Kadakia, M.P.

Notes: The authors examined transcriptional regulation of the vitamin D receptor (VDR) by p53 and p63, a member of the p53 family, under stressed and unstressed conditions. Reporter constructs with the full-length and minimal VDR promoters controlling expression of firefly luciferase were cotransfected with p53 or p63 expression constructs, and transcriptional activation of the VDR promoter was monitored using the Dual-Luciferase® Reporter 100 Assay System. Results were normalized to Renilla luciferase activity. Interaction between p73, another member of the p53 family, and the VDR promoter was examined using chromatin immunoprecipitation. The imunnopreciptated chromatin was reverse crosslinked, DNA was eluted and VDR and p21 sequences were detected by PCR using GoTaq® Green Master Mix. (3715)

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Plant Physiol. 145, 547–558. Diversity of acetyl-coenzyme A carboxylase mutations in resistant Lolium populations: Evaluation using clethodim. 2007

Yu, Q., Collavo, A., Zheng, M.Q., Owen, M., Sattin, M. and Powles, S.B.

Notes: The authors characterized mutations in the acetyl-coenzyme A carboxylase (ACCase) gene that confer resistance to the herbicide clethodim in the grass weed Lolium rigidum. The ACCase gene was amplified from clethidem-resistant and susceptible plants, then sequenced to identify previously unknown mutations. Amplifications of ACCase were performed using 300ng of genomic DNA and GoTaq® Green Master Mix. The Wizard® SV Gel and PCR Clean-Up System was used to purify PCR products directly or from agarose gels. (3721)

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Microbiology 153, 3023–3033. Expression analysis of extracellular proteins from Phanerochaete chrysosporium grown on different liquid and solid substrates. 2007

Sato, S., Liu, F., Koc, H. and Tien, M.

Notes: The authors characterized expression of extracellular proteins by white-rot fungus, Phanerochaete chrysosporium, grown on wood. Temporal expression of these proteins was monitored by relative quantitative RT-PCR. Two micrograms of total RNA was reversed transcribed using 1µg of Random Primers at 37°C for 1 hour. PCRs with one set of PCR primers were performed using 0.5 units of GoTaq® DNA Polymerase, 1X reaction buffer, 250µM each dNTP, 0.5µM each primer and 1µl of cDNA. PCRs with two sets of PCR primers were performed using 2.5 units of GoTaq® DNA Polymerase, 1.6X reaction buffer, 500µM each dNTP, 0.5µM each primer and 1µl of cDNA. (3708)

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J. Clin. Invest. 117, 3042–3048. HLA class I polymorphisms are associated with development of infectious mononucleosis upon primary EBV infection. 2007

McAulay, K.A., Higgins, C.D., Macsween, K.F., Lake, A., Jarrett, R.F., Robertson, F.L., Williams, H. and Crawford, D.H.

Notes: The authors examined whether genetic differences at the HLA class I locus affect development of Epstein Barr Virus-associated diseases. Peripheral blood mononuclear cells were isolated from asymptomatic EBV-seropositive and seronegative individuals and patients with acute infectious mononucleosis. DNA was isolated, and genotypes at two HLA class I loci and one HLA class III locus, as a control, were determined by PCR. The 10µl PCRs contained 25ng of DNA, 1X GoTaq® Flexi Reaction Buffer, 2.5mM MgCl2, 200µM dNTP, 0.5 units of GoTaq® Flexi DNA Polymerase and 25µM of forward and reverse primer, one of which was labeled with 6-FAM fluorescent dye. The results show that HLA class I polymorphisms might predispose people to develop infectious mononucleosis upon EBV infection. (3712)

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J. Clin. Microbiol. 45, 1469–1477. Multilocus sequence typing of the pathogenic fungus Aspergillus fumigatus. 2007

Bain, J.M., Tavanti, A., Davidson, A.D., Jacobsen, M.D., Shaw, D., Gow, N.A. and Odds, F.C.

Notes: The authors developed a multilocus sequence typing scheme (MLST) to examine sequence variation and discriminate between Aspergillus fumigatus strains. They also examined the distribution of MAT1-1 and MAT1-2 sexual idiomorphs in 100 clinical and environmental isolates. Sexual idiomorphs were determined using PCR and a reverse primer to both idiomorphs and a forward primer specific to either MAT-1 or MAT-2. PCRs consisted of 2mM MgCl2, 200µM DNTPs and 2.5 units of GoTaq® DNA Polymerase. (3714)

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Am. J. Pathol. 171, 1312–1323. Target genes of neuron-restrictive silencer factor are abnormally up-regulated in human myotilinopathy. 2007

Barrachina, M., Moreno, J., Juvés, S., Moreno, D., Olivé, M. and Ferrer, I.

Notes: These authors used chromatin immunoprecipitation to show that neuron-restrictive silencer factor interacts with the ubiquitin carboxy-terminal hydrolase L1 (UCHL1) promoter in U87-MG, DMS53 and HeLa cells. The neuron-restrictive silencing element (NRSE1) of the UCHL1 promoter was amplified using GoTaq® Flexi DNA Polymerase in a 25µl PCR. (3705)

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Exp. Parasitol. 112, 63-65. Babesia canis vogeli: A novel PCR for its detection in dogs in Australia. 2006

Martin, A.R., Dunstan, R.H., Roberts, T.K., and Brown, G.K.

Notes: GoTaq® DNA Polymerase was used in PCR to test dog blood for the presence of Babesia canis. Genomic DNA isolated from dog blood was analyzed with primers to the variable 5’ region of the Babesia canis 18S rRNA gene. PCR was performed in 50µl reactions containing 1.25 units of GoTaq® DNA Polymerase and 10µl of GoTaq® Reaction Buffer. (3367)

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Nucl. Acids Res. 34, 6215-6224. Chromosomal integration of LTR-flanked DNA in yeast expressing HIV-1 integrase: down regulation by RAD51 2006

Desfarages, S., San Filippo, J., Fournier, M. Calmels, C., Caumont-Sarcos, A., Litvak, S., Sung, P., Parissi, V.

Notes: In the process of demonstrating the role of IN in HIV-1 integration in yeast, the authors purified all DNA vectors and PCR products with the Wizard® Plus SV Miniprep System and Wizard® SV Gel System. PCR products were generated using Taq DNA Polymerase. The pGEM®-T Vector was used to clone amplification products. Sequencing was performed using BamHI, religated with T4 DNA Ligase. (3704)

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Pesticide Biochem. Physiol. 84, 236-242. Deletion of Cyp6d4 does not alter toxicity of insecticides to Drosophila melanogaster. 2006

Hardstone, M.C., Baker, S.A., Gao, J., Ewer, J., and Scott, J.G.

Notes: Researchers used the GoTaq® DNA Polymerase in PCR screens for excisions around a CYP6d4 gene in the HA-1829 strain of Drosophila. PCR was performed in a 20μl reaction volume using 0.4 Units of GoTaq® DNA Polymerase, 2.75mM MgCl2 and 1μl of DNA (equivalent to the DNA in approximately 1/5 to 1/10 of a fly). (3363)

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J. Appl. Microbiol. 98, 1001-1009. Detection of lactococcal 936-species bacteriophages in whey by magnetic capture hybridization PCR targeting a variable region of receptor-binding protein genes. 2006

Dupont, K., Vogensen, F.K., and Josephsen, J.

Notes: GoTaq® DNA Polymerase was used in PCR to detect Lactococcus lactis phage DNA strains in whey samples. Phage DNA templates were amplified directly from DNase treated and boiled whey samples. For these reactions, the researchers use 0.25µl (1.25 units) of GoTaq® DNA Polymerase for each 50μl reaction. Primers were designed to distinguish various strains of Lactococcus lactis phage receptor-binding protein genes. (3362)

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Vet. Parasitol. 135, 99-104. Determination of prevalence and risk factors of infection with Babesia in small ruminants from Greece by polymerase chain reaction amplification. 2006

Theodoropoulos, G., Gazouli, M., Ikonomopoulos, J.A., Kantzoura, V., and Kominakis, A.

Notes: Researchers used GoTaq® DNA Polymerase to test sheep and goat blood samples for the presence of Babesia DNA. Primers were designed around the 18S rRNA sequence of Babesia sp. PCR was performed in a 50µl reaction volume using 1 unit of GoTaq® DNA Polymerase. Ten microliters of each amplification reaction were loaded on gels and subjected to electrophoresis. (3380)

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Plant Sci. 170, 705-14. Expression profile analysis and biochemical properties of the peptide methionine sulfoxide reductase A (PMSRA) gene family in Arabidopsis. 2006

Romero, H.M., Pell, E.J. and Tien, M.

Notes: PMSRA expression was examined in 4-week old plants exposed to 10μM methyl viologen, 100μM cercosporin, photo-oxidative stress or ozone. Samples were ground in liquid nitrogen and total RNA isolated. Total RNA (2.5μg) was reverse transcribed into cDNA with random primers d(N)10, then amplified using gene-specific primers for PMSRA1—PMSRA5 and an antibody-mediated hot start containing a mixture of GoTaq® DNA Polymerase and Taq antibody (BD Biosciences, Mountain View CA). In a total volume of 25μl, the RT-PCR reaction mixture contained 2.5 units of GoTaq® DNA Polymerase, 10mM Tris–HCl (pH 8.5), 60mM KCl, 2.4mM MgCl2 and 300μM of each dNTP. For each RT-PCR, a plant 18S universal internal standard (Ambion, Austin TX) was included as a loading control. Amplification reaction conditions were as follows: 27 cycles at 94°C for 45 seconds, 55°C for 45 seconds and 72°C for 1 minute. For each analysis, three rounds of RT-PCR were conducted with three independently isolated total RNA samples. Twenty microliters from each PCR were fractionated by 1.5% (w/v) agarose gel in Tris–borate EDTA buffer and stained with 0.5% (w/v) ethidium bromide. (3370)

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BMC Genetics 7, 13. High resolution physical map of porcine chromosome 7 QTL region and comparative mapping of this region among vertebrate genomes. 2006

Demars, J., Riquet, J., Feve, K., Gautier, M., Morisson, M., Demeure, O., Renard, C., Chardon, P., and Milan, D.

Notes: Fifteen microliter amplification reactions performed using 0.5 Units of GoTaq® DNA Polymerase and 25 ng BAC or genomic DNA were used as templates for sequencing portions of the QTL region of porcine chromosome 7. Amplification products were verified by gel electrophoresis followed by ethidium bromide staining. (3361)

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J. Neurosci. 26, 3299-3308. Human astrocytes are resistant to Fas ligand and tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis. 2006

Song, J.H., Bellail, A., Tse, M.C.L., Yong, V.W. and Hao, C.

Notes: Total RNA was extracted from human astrocytes and control A549 cells. First strand cDNA was synthesized from 3μg of total RNA using random hexamers. PCR was performed on the cDNA samples using primers for DR4, DR5, and GAPDH with GoTaq® Green Master Mix. The PCR was performed with an initial denaturation at 94°C for 2 minutes, followed by cycles of 30 seconds at 95°C, 30 seconds at 55°C, and 45 seconds at 72°C. The PCR products were analyzed on a 1.5% agarose gel and stained with ethidium bromide. (3372)

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J. Biol. Chem. 281, 30001–30014. Identification of a domain in the V0 subunit d that is critical for coupling of the yeast vacuolar proton-translocating ATPase. 2006

Owegi, M.A., Pappas, D.L., Finch, M.W. Jr., Bilbo, S.A., Resendiz, C.A., Jacquemin, L.J., Warrier, A., Trombley, J.D., McCulloch, K.M., Margalef, K.L., Mertz, M.J., Storms, J.M., Damin, C.A. and Parra, K.J.

Notes: The authors generated truncations of subunit d of a vacuolar proton-translocating ATPase pump to examine the protein's function. The N- and C-termini were required for subunit stability as well as pump function and assembly. Function was restored to the C-terminal deletions of subunit d by creating a chimeric protein with the C-terminus of a subunit d homolog from Thermus thermophilus. Three tandem repeats of a hemagglutinin (HA3) epitope were amplified and inserted at the N-terminus so that the chimeric protein could be visualized by Western blot with an anti-HA antibody. Initial amplification of the HA3 epitope and amplifications to confirm the orientation of the epitope were performed using GoTaq®Green Master Mix. (3720)

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Vet. Immunol. Immunopathol. 110, 279-92. Perforin expression can define CD8 positive lymphocyte subsets in pigs allowing phenotypic and functional analysis of natural killer, cytotoxic T, natural killer T and MHC un-restricted cytotoxic T-cells. 2006

Denyer, M.S., Wileman, T.E., Stirling, C.M.A., Zuber, B., and Takamatsu, H.

Notes: In this study, GoTaq® DNA Polymerase was used in two-step RT-PCR. The ImProm-II™ Reverse Transcription System was first used to produce cDNA using an oligo d(T)15 primer. PCR was then performed using GoTaq® DNA Polymerase. Each reaction contained 2μl cDNA, 10μl GoTaq® Reaction Buffer, 1μl dNTP (10mM), 0.2μl GoTaq® DNA Polymerase, 1μl each primer (10pmol) and 34.8μl nuclease-free water. PCR was performed at 94°C for 30 seconds, 55°C for 30 seconds, 72°C for 60 seconds for 35 cycles, and 72°C for 10 minutes.PCR products were visualized by agarose gel electrophoresis containing ethidium bromide and then sequenced.
(3368)

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FEBS Lett. 579, 832-8. Phosphodiesterase inhibitors stimulate osteoclast formation via TRANCE/RANKL expression in osteoblasts: possible involvement of ERK and p38 MAPK pathways. 2006

Takami, M., Cho, E.S., Lee, S.Y., Kamijo, R. and Yim, M.

Notes: Mouse bone marrow cells and calvarial osteoblasts were cocultured for 6 days with or without 50 μM of IBMX. Total RNA was then isolated from the cells and cDNA templates prepared. cDNAs were subjected to PCR amplification with GoTaq® DNA Polymerase. Primers for mouse PDE4s, TRANCE, CTR, cathepsin K and GAPDH genes were used in this study. The PCR program was as follows: 32 (all mouse PDE4s, TRANCE, CTR, and cathepsin K) or 28 (GAPDH) cycles, after an initial denaturation step at 94°C for 3 minutes, then denaturation at 94°C for 30 seconds, annealing at 58°C for 45 seconds, and extension at 72°C for 60 seconds, with a final extension at 72°C for 10 minutes. (3356)

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Mol. Cell. Biol. 26, 39-49. Recruitment of DNA damage checkpoint proteins to damage in transcribed and nontranscribed sequences. 2006

Jiang, G. and Sancar, A.

Notes: In this study, GoTaq® DNA Polymerase was used in amplification reactions to test for the recruitment of DNA damage checkpoint proteins RPA, Rad9, and ATR in ChIP assays. DNA was isolated from cells treated with UV irradiation or acetoxyacetylaminofluorene. The DNA and DNA damage checkpoint proteins RPA, Rad9, and ATR were crosslinked together and the complexes immunoprecipitated with antibodies toward RPA, Rad9, and ATR. Amplification reactions contained primers designed to amplify regions of the p53 and β-globin genes. (3366)

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Genes Dev. 19, 77-89. A dicer-like protein in Tetrahymena has distinct functions in genome rearrangement, chromosome segregation, and meiotic prophase. 2005

Mochizuki, K. and Gorovsky, M.A.

Notes: GoTaq® DNA Polymerase was used in RT-PCR. First-strand cDNA was synthesized from 3μg of total RNA. PCR was performed using the first-strand cDNA as a template, the primers TM4SF2, and GoTaq® DNA Polymerase, with 42 cycles of 94°C for 30 seconds, 50°C for 30 seconds, and 72°C for 60 seconds. (3354)

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Virology 340, 245-254. Acute respiratory infection with mouse adenovirus type 1. 2005

Weinberg, J.B., Stempflea, G.S., Wilkinsonb, J.E., Youngerc, J.G., and Spindler, K.R.

Notes: Mouse adenovirus type 1 (MAV-1) was detected in DNA extracted from the lungs of mice after PCR amplification of the E1A region of MAV-1. For these assays, 80ng of total DNA was added to a 20µl PCR reaction containing 0.5 units of GoTaq® DNA Polymerase, 4µl of 5X GoTaq® Buffer, dNTPs and primers for MAV-1 E1A. The amplified products were separated on a 1.8% agarose gel and stained with ethidium bromide. (3381)

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