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Nucl. Acids Res. 37, 5343–5352. Qri7/OSGEPL, the mitochondrial version of the universal Kae1/YgjD protein, is essential for mitochondrial genome maintenance. 2009

Oberto, J., Breuil, N., Hecker, A., Farina, F., Brochier-Armanet, C., Culetto, E. and Forterre, P.

Notes: In this paper, the role of the KAE1/osgep/ygjD gene family, a universally conserved gene set without an assigned function, was investigated in yeast and Caenorhabditis elegans. Genomic DNA was isolated from Saccharomyces cerevisiae using the Wizard® SV Genomic DNA Purification System. This purified DNA was then used in PCR. (4065)

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Nucl. Acids Res. 37, 7468–7482. Enhanced gene repair mediated by methyl-CpG-modified single-stranded oligonucleotides. 2009

Bertoni, C., Rustagi, A. and Rando, T.A.

Notes: This paper explored the effect of methyl-CpGmodified single-stranded oligonucleotides (ssODN) on gene repair. The Wizard® Genomic DNA Purification Kit was used to isolate gDNA from methylCpG-ssODN-treated myoblasts derived from limb muscle of neonatal C57 mice that stably expressed the mismatch repair site. One microgram of purified gDNA was digested overnight with 5U of restriction enzyme, purified, resuspended in 20µl of water and 5µl used in real-time PCR to determine if the target had been repaired. (4062)

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J. Biol. Chem. 284, 3682–3690. Human flap endonuclease I is in complex with telomerase and is required for telomerase-mediated telomere maintenance. 2009

Sampathi, S., Bhusari, A., Shen, B. and Chai, W.

Notes: The authors explored the role of a DNA replication factor, flap endonuclease I (FEN1), in regulating telomerase activity in mammalian cells. PCR was used to add a myc tag to the N terminus of FEN1 cDNA. The amplimer was gel purified, digested with NheI and SmaI, and cloned into the same sites in the pCI-neo Mammalian Expression Vector. The insert was confirmed by sequencing. (4030)

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Appl. Environ. Microbiol. 75, 5600–6. Single nucleotide polymorphism-based diagnostic system for crop-associated Sclerotinia species. 2009

Andrew, M. and Kohn, L.M.

Notes: The authors developed a single nucleotide polymorphism (SNP)-based assay to distinguish four Sclerotinia species. The assay consisted of amplification of a 300bp intergenic spacer and portions of the calmodulin and ras genes, followed by Southern blot using species-specific, radiolabeled probes. Amplifications were performed using the GoTaq® Colorless Master Mix, 0.2µM of each primer and 10–20ng of template DNA. (4098)

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J. Gen. Virol. 90, 1461–1470. Murid herpesvirus-4 lacking thymidine kinase reveals route-dependent requirements for host colonization. 2009

Gill, M.B., Wright, D.E., Smith, C.M., May, J.S. and Stevenson, P.G.

Notes: The authors examined the role of thymidine kinase (TK) in establishing a herpesvirus infection via the upper respiratory tract. DNA was purified from ex vivo organs of female BALB/c mice infected with a murid herpesvirus-4 (MuHV-4) TK knockout using the Wizard® Genomic DNA Purification Kit. Real-time PCR was used with 50–80ng of purified DNA to determine viral load of the animals. (4015)

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Plant Physiol. 150, 1356–1367. Sucrose control of translation mediated by an upstream open reading frame-encoded peptide. 2009

Rahmani, F., Hummel, M., Schuurmans, J., Wiese-Klinkenberg, A., Smeekens, S. and Hanson, J.

Notes: The authors were wanted to study the upstream open reading frame 2 (uORF2) of the 5’ leader of bZIP11 mRNA, which has a role in sucrose regulation. The whole 5’ leader fragment of bZIP11 was subcloned into the pALTER® Vector and amino acid substitutions were introduced using the Altered Sites® II in vitro Mutagenesis System. The pGEM®-T Easy Vector was used to clone two PCR fragments that were then subcloned using restriction enzymes to create a fusion of uORF2 to a different 5’ leader. Arabidopsis seedlings were transformed via particle bombardment. 20mg of plant tissue was ground in Passive Lysis Buffer, centrifuged, and 20µl of the supernatant was assessed for reporter gene expression using the Dual-Luciferase® Reporter Assay System. (4023)

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Food Control [epub ahead of print]. Evaluation of DNA extraction procedures for traceability of various tomato products. 2009

Turci, M., Sardaro, M.L.S., Visioli, G., Maestri, E., Marmiroli, M. and Marmiroli, N.

Notes: In this study, the authors wanted to examine the ability to trace the origin of tomato goods from fresh to processed. They tested several DNA extraction procedures for fresh tomato, tomato sauce, tomato puree, tomato pulp, whole peeled S. Marzano PDO (Protected Designation of Origin) tomato, whole peeled tomato, tomato concentrate and ‘‘Arrabbiata sauce”. Homogenized material (200mg) was extracted in three replicates using seven different methods including the Wizard® DNA Clean-Up System. The DNA extracted was then analyzed by agarose gel electrophoresis, quantified and tested in PCR using SSR loci. The authors concluded that the Wizard® DNA Clean-Up System was the most effective of the DNA extraction methods tested and yielded the greatest number of successful amplification reactions with lowest investment of personnel time and money. (4003)

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Arterioscler. Thromb. Vasc. Biol. Sept 24, [Epub ahead of print]. Eotaxin increases monolayer permeability of human coronary artery endothelial cells. 2009

Jamaluddin, M.S., Wang, X., Wang, H., Rafael, C., Yao, Q. and Chen, C.

Notes: Glutathione levels were assessed in human coronary artery endothelial cells (HCAECs) as a measure of oxidative stress. HCAECs were treated with either 100ng/ml eotaxin, a newly discovered chemokine, or pretreated with 2µmol/l MnTBAP for 30 minutes followed by eotaxin treatment for 45 minutes. Positive controls were treatment with 10µg/ml antimycin A and 2ng/ml TNF-α. Cellular glutathione was measured using the GSH-Glo™ Glutathione Assay. (4011)

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Infect. Immun. 77, 3234–43. Sab, a novel autotransporter of locus of enterocyte effacement-negative shiga-toxigenic Escherichia coli O113:H21, contributes to adherence and biofilm formation. 2009

Herold, S., Paton, J.C. and Paton, A.W.

Notes: To try to understand the mechanism by which certain strains of Shiga-toxigenic E.coli adhere to host intestinal epithelium, the authors characterized the novel autotransporter protein Sab. Expression levels and protein localization were examined by Western blot analysis and enzyme-linked immunosorbent assay. The mouse anti-Sab antibody used in these studies was raised against an N-terminal His6-Sab fusion protein purified using the HisLink™ Resin. (4102)

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J. Nutr. 139, 1054–1060. Folic acid supplementation during the juvenile-pubertal period in rats modifies the phenotype and epigenotype induced by prenatal nutrition. 2009

Burdge, G.C., Lillycrop, K.A., Phillips, E.S., Slater-Jefferies, J.L., Jackson, A.A. and Hanson, M.A.

Notes: This study examined the effects of folic acid supplementation on the offspring of pregnant rats fed a protein-restricted diet. Genomic DNA was extracted from rat adipose tissue using the Wizard® SV Genomic DNA Purification System. The purified DNA was then incubated with methylation-sensitive restriction enzymes and then used in real-time PCR. (4066)

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Clin. Can. Res. 15, 2523–2530. Identification of CD20 C-terminal deletion mutations associated with loss of CD20 expression in non-Hodgkin's lymphoma. 2009

Terui, Y., Mishima, Y., Sugimura, N., Kojima, K., Sakurai, T., Mishima, Y., Kuniyoshi, R., Taniyama, A., Yokoyama, M., Sakajiri, S., Takeuchi, K., Watanabe, C., Takahashi, S., Ito, Y. and Hatake, K.

Notes: The researchers examined if a CD20 mutation would affect resistance to rituximab, an adjunct cancer therapy drug used for CD20-positive B-cell lymphoma. CD20 PCR products amplified from genomic DNA were cloned into the pTARGET™ Mammalian Expression Vector. These CD20 mutant constructs were stably introduced into K562 chronic myelogenous leukemia cells by electroporation and selected using G-418. One microgram of CD20 mutant construct DNA was transcribed and translated using an in vitro translation kit from Promega. (4032)

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Nucl. Acids Res. 37, 2070–86. HMGB1 and HMGB2 proteins up-regulate cellular expression of human topoisomerase IIα. 2009

Stros, M., Polanská, E., Struncová, S. and Pospísilová, S.

Notes: The authors examined whether HMGB1 and HMGB2 proteins could affect promoter activity of the topoisomerase IIα gene. Portions of the topoisomerase IIα gene promoter were cloned into the pGL3 Basic Vector, and Saos-2 cells were cotransfected with the resulting constructs, an HMGB1- or HMGB2-expressing plasmid and the pRL-tk Vector as a control for normalization. Firefly and Renilla luciferase activities were determined using the Dual-Luciferase Reporter Assay. To determine whether HMGB1 and HMGB2 promoted binding of the transcription factor nuclear factor-Y (NF-Y) to the topoisomerase IIα promoter, the authors used a chromatin immunoprecipitation (ChIP) assay. Two populations of Saos-2 cells, one of which expressed HMGB1 or HMGB2 and one that had expression inhibited, were fixed with formaldehyde, then treated to shear chromatin. Immunoprecipitation was performed using an anti-NF-Y antibody, and the amount of DNA bound to the NF-Y was quantified by semi-quantitative PCR using GoTaq® Hot Start DNA Polymerase and Green GoTaq® Flexi Reaction Buffer. (4037)

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Proc. Natl. Acad. Sci. USA 106(23), 9262-7. Long-lived Indy and calorie restriction interact to extend life span. 2009

Wang, P.Y., Neretti, N., Whitaker, R., Hosier, S., Chang, C., Lu, D., Rogina, B., and Helfand, S.L.

Notes: These authors investigated the relationship between calorie restriction and INDY expression on lifespan in Drosophila melanogaster. They showed that calorie restriction downregulates INDY expression in normal flies. INDY mutants on a normal diet had increased lifespan that was not extended further by calorie restriction. The INDY long-lived flies also shared several phenotypic characteristics with normal flies on a calorie-restricted diet. The authors then demonstrated that the phenotypic effects of the INDY mutation were not caused by reduced ability to intake food. The results show that INDY and calorie restriction interact to extend lifespan, and that decreased INDY expression induces a calorie-restriction-like status. During the study, the Maxwell 16 Tissue DNA Purification System was used to isolate DNA from Drosophila. The DNA was used in PCR analyses to confirm the absence of Wolbachia DNA in the experimental lines. (4001)

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J. Exp. Bot. 60, 1409-25. A strong effect of growth medium and organ type on the identification of QTLs for phytate and mineral concentrations in three Arabidopsis thaliana RIL populations. 2009

Ghandilyan, A., Ilk, N., Hanhart, C., Mbengue, M., Barboza, L., Schat, H., Koornneef, M., El-Lithy, M., Vreugdenhil, D., Reymond, M. and Aarts, M.G.

Notes: Mineral accumulation was studied in Arabidopsis thaliana comparing loci involved with growing in soil versus hydroponics. An F2 population derived from a cross between Landsberg erecta (Ler; maternal parent) and Eringsboda-1 (Eri-1; paternal parent) was propagated by single seed descent for nine successive generations in soil.
The flower buds of three plants per line were collected, and the DNA extracted using the Wizard® Magnetic 96 DNA Plant System and used for genotyping with 90 amplified fragment length polymorphism PCR (AFLP) and 39 single sequence length polymorphisms (SSLP) markers to build a genetic map of quantitative trait loci (QTL). (4136)

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Forensic Sci. Int. Genet. 3, 14–21. Developmental validation of a real-time PCR assay for the simultaneous quantification of total human and male DNA. 2008

Krenke, B.E., Nassif, N., Sprecher, C.J., Knox, C., Schwandt, M. and Storts, D.R.

Notes: The authors describe the developmental validation of the Plexor® HY System, a quantitative PCR assay that simultaneously quantifies total human and male DNA. Validation studies examined: (1) human specificity, (2) sensitivity, (3) quantitation of degraded DNA, (4) impact of inhibitors, (5) male/female mixture and Y-assay male specificity, (6) reproducibility and concordance and (7) population studies. (3969)

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J. Biol. Chem. 283, 16868–75. The hairless phenotype of the Hirosaki hairless rat is due to the deletion of an 80-kb genomic DNA containing five basic keratin genes. 2008

Nanashima, N., Akita, M., Yamada, T., Shimizu, T., Nakano, H., Fan, Y. and Tsuchida, S.

Notes: The mutation responsible for the hairless phenotype was linked to a 80kb deletion of chromosome 7q36. Because many basic keratin genes are located at 7q36, the authors examined keratin gene expression in the Hirosaki hairless rat using RT-PCR. Expression of kb21, kb23 and kb26 was not detected, whereas other basic keratin genes were expressed. RT-PCR was performed using the AccessQuick™ RT-PCR System and 0.5µg of total RNA isolated from rat skin for 21–30 cycles. (3887)

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Genetics 178, 1415–29. Comparative genetics of sex determination: masculinizing mutations in Caenorhabditis briggsae. 2008

Kelleher, D.F., de Carvalho, C.E., Doty, A.V., Layton, M., Cheng, A.T., Mathies, L.D., Pilgrim, D. and Haag, E.S.

Notes: The authors characterized masculinizing mutations of the female-promoting tra genes in Caenorhabditis briggsae (Cb-tra). Using RT-PCR, the authors monitored the levels of full-length Cb-tra mRNA and a novel splice variant; actin mRNA was amplified as a control. RT-PCR was carried out using the AccessQuick™ RT-PCR System and RNA from 5–10 worms per 50µl reaction. (3892)

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Appl. Environ. Microbiol. 74, 1886–91. Use of Drosophila S2 cells as a model for studying Ehrlichia chaffeensis infections. 2008

Luce-Fedrow. A., Von Ohlen, T., Boyle, D., Ganta, R.R. and Chapes, S.K.

Notes: The authors infected Drosophila S2 cells with Ehrlichia chaffeensis to determine if Drosophila is a model system to study E. chaffeensis pathogenesis. E. chaffeensis was also grown in canine macrophage-like DH82 cells, the most common host cell line. Infections were assessed by RT-PCR using the Access RT-PCR System, 0.5–1.0µg of RNA and primers specific to the E. chaffeensis 16S rRNA gene. Housekeeping genes for ribosomal protein 49 and canine glyceraldehyde-3-phosphate dehydrogenase were amplified as control targets for Drosophila and DH82 cells, respectively. Negative controls without reverse transcriptase were performed to be sure that DNA was absent from the RNA samples. (3888)

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J. Cell Sci. 121, 504–13. Integrin alpha9 beta1 is a receptor for nerve growth factor and other neurotrophins. 2008

Staniszewska, I., Sariyer, I.K., Lecht, S., Brown, M.C., Walsh, E.M., Tuszynski, G.P., Safak, M., Lazarovici, P. and Marcinkiewicz, C.

Notes: The authors investigated the ability of α9β1 integrin to act as a neurotrophin receptor and affect cell signaling pathways. As part of the study, RT-PCR was used to detect the presence of other neurotrophin receptors in their model cell line, SW480. Reverse transcription was performed using the Reverse Transcription System and 1µg of total RNA isolated using the SV Total RNA Isolation System. The resulting cDNA (5µg) was amplified for 35 cycles (β-actin as a control) or 40 cycles (TrkA and p75NTR) using GoTaq® Green Master Mix. RT-PCR results were confirmed by Western blot analysis. (3884)

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Proc. Natl. Acad. Sci. USA 105, 8914-8919. An epoxide hydrolase involved in the biosynthesis of an insect sex attractant and its use to localize the production site. 2008

Abdel-Latief, M., Garbe, L.A., Koch, M., and Ruther, J.

Notes: These authors amplified and characterized a putative epoxide hydrolase gene from the jewel wasp Nasonia vitripennis. PCR fragments were amplified from genomic DNA, purified from gels using the Wizard® SV Gel and PCR Clean Up System and then subcloned into the pGEM®-T Easy Vector. The plasmid DNA was purified using the PureYield™ Midiprep System. Linearized plasmids were used for in vitro transcription of RNA for use in RNA interference experiments. (3903)

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J. Clin. Microbiol. 46, 3752–3758. Plastic polymers for efficient DNA microarray hybridization: application to microbiological diagnostics. 2008

Zhao, Z., Peytavi, R., Diaz-Quijada, G.A., Picard, F.J., Huletsky, A., Leblanc, E., Frenette, J., Boivin, G., Veres, T., Dumoulin, M.M. and Bergeron, M.G.

Notes: A plastic support suitable for use in microfluidic systems for highly sensitive DNA microarray hybridizations was developed and tested. Human DNA from Hsap-11 cells was isolated using the MagneSil® KF, Genomic System on a KingFisher ML instrument. Ten nanograms of the isolated DNA was used in RT-PCR. (4021)

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Genetics 179, 177-192. The small ubiquitin-like modifier (SUMO) and SUMO-conjugating system of Chlamydomonas reinhardtii. 2008

Wang, Y., Ladunga, I., Miller, A.R., Horken, K.M., Plucinak, T., Weeks, D.P. and Bailey, C.P.

Notes: These authors used computational biology to screen the genome of the alga Chlamydomonas reinhardtii for SUMO (small ubiquitin-like modifier) homologs. They identified several SUMO and SUMO-like sequences. One of these proteins, crSUMO96, which was recognized by the A. thaliana anti-SUMO antibody, was studied in detail. During their studies, the authors used the PureYield™ RNA Midiprep System to isolate total RNA from C. reinhardtii cells. This RNA was used in real-time RT-RCR assays to detect mRNA transcripts for the various SUMO-like proteins. The Plexor® Two-Step qRT-PCR System was used for the real-time assays. For expression studies, cDNA encoding the various proteins was amplified and subcloned into the pGEM®-T Easy Vector before transfer into an expression vector. (3875)

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Endocrinology 149, 2306–12. The gastrointestinal hormone ghrelin modulates inhibitory neurotransmission in deep laminae of mouse spinal cord dorsal horn. 2008

Vergnano, A.M., Ferrini, F., Salio, C., Lossi, L., Baratta, M. and Merighi, A.

Notes: The authors investigated the expression pattern of type 1a growth hormone secretagogue receptor (type 1a GHSR), a receptor for ghrelin. In situ RT-PCR was performed on paraformaldehyde-fixed, paraffin-embedded mouse spinal cord tissue. Prior to in situ RT-PCR, tissue sections were treated with proteinase K and triethanolamine, then dewaxed. Reverse transcription was performed using the Reverse Transcription System and oligo (dT)15 primers; followed by amplification using the PCR Master Mix in the presence of 11-digoxygenin-dUTP (1mM). Amplification products were detected using an anti-digoxygenin, alkaline phosphatase-conjugated goat antibody and nitro blue tetrazolium/5-bromo-3-indolylphosphate-p-toluidine salt (NBT/BCIP). (3968)

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J. Biol. Chem. 283, 23514–23. Snail regulates cell-matrix adhesion by regulation of the expression of integrins and basement membrane proteins. 2008

Haraguchi, M., Okubo, T., Miyashita, Y., Miyamoto, Y., Hayashi, M., Crotti, T.N., McHugh, K.P. and Ozawa, M.

Notes: Snail is a transcriptional repressor of E-cadherin that enhances both cell attachment and cell detachment in Madin Darby canine kidney (MDCK) and A4231 cells. To investigate this effect, the authors used Western blot analysis and RT-PCR to monitor protein and mRNA levels of the major adhesive proteins expressed in epithelial cells: laminin, heparin sulfate proteoglycan and collagens. For RT-PCR, total RNA was isolated from transiently transfected snail-expressing MDCK and A431 cells and untransfected cells, then reverse transcribed. The resulting cDNA was amplified by PCR using GoTaq® DNA Polymerase; glyceraldehyde-3-phosphate dehydrogenase was amplified as an internal control. The ability of Snail to regulate the integrin αV promoter was also examined by cloning the promoter and several promoter deletions upstream of a firefly luciferase reporter gene in the pGL3-Basic Vector. Each of these constructs (1µg) and 20ng of pRL-CMV Vector were transfected into MDCK and MDCK/snail cells, and luminescence was measured using the Dual Luciferase Assay System. (3882)

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Mol. Cancer Res. 6, 546–54. CXCL16 functions as a novel chemotactic factor for prostate cancer cells in vitro. 2008

Lu, Y., Wang, J., Xu, Y., Koch, A.E., Cai, Z., Chen, X., Galson, D.L., Taichman, R.S. and Zhang, J.

Notes: The authors sought to detect expression of the chemokine CXCL16 and its receptor, CXCR6, in prostate cancer cell lines and in benign and malignant prostate cancer tissues. The Access RT-PCR System was used to amplify and detect CXCL16 and CXCR6 mRNA in these cells and tissues. Each RT-PCR contained 1µg of total RNA, and amplifications were carried out for 35 cycles. Amplified products were detected on a 1.5% ethidium bromide-stained agarose gel. (3836)

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