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Methods 67, 36–44. Construction of Specific Parallel Amplification of RNA Ends (SPARE) libraries for the systematic identification of plant microRNA processing intermediates 2014

Schapire, A.L., Bologna, N.G., Moro, B., Zhai, J., Meyers, B.C. and Palatnik, J.F.

Notes: The authors of this study present a method for genome-wide analysis of miRNA processing intermediates in plants. The method presented uses reverse transcription performed with a mix of primers designed against known miRNA precursors. The molecules are next amplified to generate a library for deep sequencing. Total RNA was isolated and purified from plant tissue and then treated with RQ1 RNase-Free DNase to remove contaminating DNA. Ribosomal RNA was selectively removed before adapter ligation and reverse transcription. cDNA libraries were prepared; PCR products from the libraries were separated on an agarose gel and purified using Wizard® SV Gel and PCR Clean-Up System prior to sequencing. (4561)

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Soil Biol. Biochem. 64, 18–27. Contrasting Euryarchaeota communities between upland and paddy soils exhibited similar pH-impacted biogeographic patterns 2013

Wu, H-W., Zhang, L-M, Yuan, C-L. and He, J-Z.

Notes: Products from PCR of the archaeal 16SrRNA gene were purified using the Wizard® SV Gel and PCR Clean-Up Kit before pyrosequencing. (4549)

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PLos ONE 8, e67151. First transcriptome and digital gene expression analysis in Neuroptera with an emphasis on chemoreception genes in Chrysopa pallens (Rambur) 2013

Li, Z.Q., Zhang, S., Ma, Y., Luo, J.Y., Wang, C.Y., Lv, L.M., Dong, S.L. and Cui, J.J.

Notes: RNA was isolated from adult C. pallens (insect) tissues using the SV Total RNA Isolation System prior to cDNA library construction and sequencing on the Illumina HiSeq™ platform. To validate NGS sequence alignment, end-to-end PCR was performed. PCR products were purified using the Wizard® SV Gel and PCR Clean-Up System prior to cloning into a T vector. To verify the identified differentially expressed genes, quantitative real-time PCR (RT-qPCR) was used. Total RNA was extracted and cDNAs synthesized using the Reverse Transcription System prior to qPCR. (4560)

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J. Biotechnol. 166, 42–50. Molecular characterization of the first transgenic common bean immune to the Bean golden mosaic virus 2013

Aragão, F.J., Noqueira, E.O. Tinoco, M.F. and Faria, J.C.

Notes: PCR products from a tertiary TAIL-PCR were separated by agarose gel electrophoresis and purified using the Wizard® SV Gel and PCR Clean-Up Kit. Purified fragments were cloned into pGEM®-T Easy Vectors, and clones were sequenced using next-generation sequencing technologies and data analyzed using the Roche GS FLX De Novo Assembler. (4547)

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Genome Biol. Evol. 5, 1298–1308. Substantial variation in the extent of mitochondrial genome fragmentation among blood-sucking lice of mammals. 2013

Jiang, H., Barker, S.C. and Shao, R.

Notes: PCR products were analyzed on a 1% agarose gel and sized with molecular markers before selection for purification using the Wizard® SV Gel and PCR Clean-Up System. The chosen purified PCR amplicons were then used in NGS. (4557)

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New Phytol. 195, 844–56. Seasonal trends in the biomass and structure of the bryophyte-associated fungal communities explored by 454 pyrosequencing 2012

Davey, M.L., Heegaard, E., Halvorsen, R., Ohlson, M. and Kauserud, H.

Notes: Genomic DNA was extracted from shoot fragments using an organic extraction procedure and purified using the Wizard® SV Gel and PCR Clean-Up System prior to template preparation by nested PCR. Products from the second PCR were separated by electrophoresis and purified using the Wizard® SV Gel and PCR Clean-Up System prior to pyrosequencing. (4548)

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PLos ONE 6, e25263. Analysis of 16S rRNA amplicon sequencing options on the Roche/454 next-generation titanium sequencing platform 2011

Tamaki, H., Wright, C.L., Li, X., Lin, Q., Hwang, C., Wang, S., Timmapuram, J., Kamagata, Y. and Liu, W.T.

Notes: DNA was isolated from a variety of environmental samples including surface soil, drinking water biofilm, sludge from an anerobic digester, bioreactor samples, ground water, peat soil and glacial deposit soil. The 16S rRNA gene was amplified from the DNA. PCR amplifications were run on agarose gels, and bands of the predicted sizes excised and purified using the Wizard® SV Gel and PCR Clean-Up System before pooling for pyrosequencing. (4554)

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Bioinformatics 27, 2027-2030. Pathogen detection using short-RNA deep sequencing subtraction and assembly 2011

Isakov, O., Modai, S. and Shomron, N.

Notes: To confirm the  Mycoplasma contamination in HIV-1 infected samples detected by high-throughput sequencing, a PCR-based Mycoplasma test was performed. Products were separated on an agarose gel and purified using the Wizard® SV Gel and PCR Clean-Up System before confirmatory sequencing. (4537)

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PLos ONE 6, e29604. Probing the SELEX process with next-generation sequencing 2011

Schütze, T., Wilhelm, B., Greiner, N., Braun, H., Franziska, P., Möri, M., Erdman, V.A., Lehrach, H., Konthur, Z., Menger, M., Arndt, P.F. and Glökler, J.

Notes: Wizard® SV Gel and PCR Clean-Up System was used to purify products after barcodes were attached by PCR prior to sequencing on an Illumina Genome Analyzer GA2. (4553)

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DNA Research 18, 93–105. The complete chloroplast genome of 17 individuals of pest species Jacobaea vulgaris: SNPs, microsatellites and barcoding markers for population and phylogenetic studies 2011

Doorduin, L., Gravendeel, B., Lammers, Y., Ariyurek, Y., Chin-A-Woeng, T. and Vrieling, K.

Notes: Wizard® SV Gel and PCR Clean-Up System was used to extract and purify DNA fragments that had been amplified by long-range PCR and separated on an agarose gel prior to next-generation sequencing on an Illumina platform. (4535)

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Nucl. Acids Res. 38, 522–33. An integrated pipeline for next-generation sequencing and annotation of mitochondrial genomes 2010

Jex, A.R., Hall, R.S., Littlewood, D.T. and Gasser, R.B.

Notes: Wizard® SV Gel and PCR Clean-Up System was used to clean up genomic DNA isolated from parasitic nematodes isolated from a variety of animals. Species identification of each nematode specimen was determined via PCR amplification of specific nuclear DNA followed by purification of the amplified product using Wizard® PCR Preps DNA Purification System before sequencing using BigDye chemistry. Wizard® SV Gel and PCR Clean-Up System was also used to prepare amplicons generated by long-PCR of mt genomes from the nematodes before NGS sequencing. Results from NGS were confirmed using PCR-based sequencing of short mt DNA tracts. Short mtDNA regions were amplified by conventional PCR. Amplicons were purified using Wizard® PCR Preps DNA Purification System before sequencing using BigDye chemistry. (4533)

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J. Biochem. 148, 721–32. A recombinant catalytic domain of matriptase induces detachment and apoptosis of small-intestinal epithelial IEC-6 cells cultured on laminin-coated surface. 2010

Mochida, S., Tsuzuki, S., Inouye, K. and Fushiki, T.

Notes: The authors determined that a recombinant catalytic domain of rat matriptase (His6t-S-CD) caused detachment of small-intestinal epithelial cells (IEC-6 cells) from laminin-coated plates. His6t-S-CD was expressed in the yeast P. pastoris and purified by ammonium sulfate precipitation, gel filtration through a PD-10 column, then Ni2+-chelating chromatography using HisLink™ Resin. The authors also treated IEC-6 cells with purified His6t-S-CD to determine if this domain induced apoptosis by monitoring annexin-V staining, DNA fragmentation and caspase-3 activity. For the DNA fragmentation analysis, IEC-6 cells were treated with His6t-S-CD, then harvested, and genomic DNA was purified using the Wizard® SV Gel and PCR Clean-Up System. DNA fragmentation was assessed by agarose gel electrophoresis and ethidium bromide staining. (4100)

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J. Biomol. Scr. 15, 418–26. Epitope mapping of antibodies using a cell array-based polypeptide library. 2010

Maier, R.H., Maier, C.J., Rid, R., Hintner, H., Bauer, J.W. and Onder, K.

Notes: The authors developed a high-density protein array using a recombinant peptide library to map the epitope recognized by a commercially available anti-vitamin D receptor (VDR) monoclonal antibody. By screening 2304 overlapping VDR peptides, they were able to identify the 37-amino-acid epitope. The library was created by amplifying the 1.2kb VDR coding region, cleaning the PCR product with the Wizard® SV Gel and PCR Clean-Up System, sonicating the PCR product, then cloning the VDR fragments into a bacterial expression vector that confers a glutathione-S-transferase (GST) tag. The epitope was verified by showing that the 37-amino-acid sequence was recognized in Western blot analysis and enzyme-linked immunosorbent assay (ELISA); the full-length VDR, also expressed as GST fusion protein, was used as a positive control. These GST fusion proteins were purified using the MagneGST™ Protein Purification System. (4154)

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Methods in Mol. Biol. 577, 25-39. High-Throughput Construction of ORF Clones for Production of the Recombinant Proteins 2009

Yamakawa, Hisashi

Notes: The authors use the Flexi® Cloning System to convert their cDNA clones to expression-ready clones. They wanted clones that could be used for comprehensive analysis with the HaloTag® Technology. They also describe a method of transfereing ORFs between Flexi® Vectors in a 96-well plate format. They also used Wizard® SV 96 Plasmid DNA Purification, Wizard® SV PCR Clean-Up, and Wizard® SV Gel and PCR Clean-Up Systems. (4056)

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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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BMC Genomics 9, 315. The complete mitochondrial genome of the Antarctic springtail Cryptopygus antarcticus (Hexapoda: Collembola). 2008

Carapelli, A., Comandi, S., Convey, P., Nardi, F. and Frati, F.

Notes: To sequence the mitochondrial genome one of the most widespread and common collembolan species of Antarctica, springtail Cryptopygus antarcticus. Specimens were collected from Killingbeck I during a 2002 polar expedition and frozen in liquid nitrogen. The Wizard® SV Genomic Purification System was used to extract total DNA from the samples and the complete mitochondrial genome was amplified twice, first with universal primers and sequenced, and then using long PCR with specific primers. The long PCR products were mechanically sheared, blunt end repaired and purified using the Wizard® SV Gel and PCR Clean-Up System. The fragments were then cloned, transformed and sequenced. (3976)

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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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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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Emerging Infect. Dis. 13, 1756-1758. Human Bocavirus infection in children with gastroenteritis, Brazil. 2007

Albuquerque, M.C., Rocha, L.N., Benati, F.J., Soares, C.C., Maranhão, A.G., Ramírez, M.L, Erdman, D., and Santos, N.

Notes: In this study, the Wizard® Genomic DNA Purification Kit was used to extract DNA from diluted fecal samples. The extracted DNA was used in PCR with specific primers to detect viral sequences. PCR fragments were gel-purified using the SV Gel and PCR Clean-Up System prior to sequencing to confirm the Bocavirus DNA identity. (4221)

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Brain Res. 1127, 66–75. Molecular characterization and gene expression of the pituitary adenylate cyclase-activating polypeptide (PACAP) in the lizard brain. 2007

Valiante, S., Prisco, M., Capaldo, A., Zambrano, I., De Falco, M., Andreuccetti, P., Laforgia, V., and Varano, L.

Notes: The authors cloned pituitary adenylate cyclase-activating polypeptide (PACAP) from lizard (Podarcis sicula) brain. They then isolated total RNA from lizard brain using the SV Total RNA Isolation System and used 4µg of total RNA in a reverse transcription with ImProm-II™ Reverse Transcriptase and oligo(dT)15 primers at 37°C for 1.5 hours. The PACAP cDNA was amplified by PCR, and the resulting PCR products were cleaned up using the Wizard® SV Gel and PCR Clean-Up System prior to sequencing. (3666)

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J. Endocrinol. 197, 201–12. Neonatal exposure to bisphenol A modifies the abundance of estrogen receptor alpha transcripts with alternative 5'-untranslated regions in the female rat preoptic area. 2007

Monje, L., Varayoud, J., Luque, E.H. and Ramos, J.G.

Notes: The authors investigated the effect of neonatal bisphenol A (BPA) exposure in rats on expression of estrogen receptor α (ERα) transcripts. Alternative ERα transcripts in preoptic area of treated and untreated rats were quantified using real-time RT-PCR. Reverse transcription was performed using 4µg of total RNA, 200pmol random primers and 300 units M-MLV Reverse Transcriptase. Real-time PCR was performed using SYBR® Green I to quantify amplified products. To determine if the changes in BPA-induced ERα transcript expression were caused by DNA methylation, the methylation status of the five ERα promoters was examined by bisulfite modification. Genomic DNA was isolated from rat tissue using the Wizard® Genomic DNA Purification Kit, denatured with NaOH, then treated with hydroquinone and sodium bisulfite. Prior to methylation-specific PCR, DNA was cleaned up using the Wizard® DNA Purification Resin as directed by the manufacturer. PCR products were cleaned up again using the Wizard® SV Gel and PCR Clean-Up System, then subjected to restriction enzyme digestion and agarose gel electrophoresis to reveal methylation-dependent sequence differences. (3911)

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J. Biol. Chem. 282, 14194-14204. Regulation of the interleukin-7 receptor α-promoter by the Ets transcription factors PU.1 and GA-binding protein in developing B cells. 2007

Dekoter, R.P., Schweitzer, B.L., Kamath, M.B., Jones, D., Tagoh, H., Bonifer, C., Hildeman, D.A., and Huang, K.J.

Notes: The interleukin-7 receptor is composed of γ and α subunits, encoded by the genes il7rg and il7r, respectively. The α subunit is expressed in developing B cells and is downregulated upon maturation. These authors investigated the mechanisms of transcriptional regulation of the il7r gene using 5´ RACE, EMSA, RNA interference and chromatin immunoprecipitation analyses. Potential promoter regions identified by 5´ RACE analysis were cloned into the pGL3-Basic luciferase reporter vector for further study. The promoter constructs were transiently transfected into the 38B9 pro-B cell line along with the control pRL-TK Vector, which expresses Renilla luciferase, and the Dual-Luciferase® Reporter Assay System was used to assess luciferase activity from the various promoter constructs. The promoter construct having the highest activity was chosen, and site directed mutagenesis was used to identify specific regions within the promoter fragment that may be important for activity. Sequence analysis was then used to identify a conserved Ets transcription factor binding site within the putative il7r promoter region. To determine whether the ETS transcription factor GABP binds to this Ets region, the authors performed chromatin immunoprecipitation analysis with an anti-GABP antibody. Immunoprecipitated DNA was then PCR-amplified with primers specific for the Ets region or control primers. The Wizard® SV Gel and PCR Clean-Up System was used to purify the amplified fragments prior to semiquantitative PCR analysis. (3626)

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Genetics 175, 1047-1058. Single-gene detection and karyotyping using small-target fluorescence in situ hybridization on maize somatic chromosomes. 2007

Lamb, J.C., Danilova, T., Bauer, M.J., Meyer, J.M., Holland, J.J., Jensen, M.D., and Birchler, J.A.

Notes: These authors generated a set of probes that could be used in fluorescence in situ hybridization (FISH) analyses for karyotyping studies on maize chromosomes. Specific target regions composed of genes or gene clusters and free from repetative elements were identified for each chromosome. Target regions were amplified by PCR, gel purified using the Wizard® SV Gel and PCR Clean-Up System, and tested in a FISH assay. Probes showing low background were selected, subcloned into the pGEM® -T Vector and sequenced to confirm identity. (3627)

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Appl. Environ. Microbiol. 73, 2860-2870. The microbial community structure in petroleum-contaminated sediments corresponds to geophysical signatures. 2007

Allen, J.P., Atekwana, E.A., Atekwana, E.A., Duris, J.W., Werkema, D.D., and Rossbach, S.

Notes: These authors studied microbial community structure at various locations in an aged underground petroleum plume. DNA was purified from soil samples collected from different sites within a contaminated area. 16S rRNA genes were then amplified from the isolated DNA, and the PCR products were run on a gel and purified using the Wizard® SV Gel and PCR Clean-Up System. After subcloning into a TA vector, the 16S RNA genes were sequenced and used to identify the various Phyla represented and characterize the microbial populations present throughout the site. (3625)

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Appl. Environ. Microbiol. 72, 6070-6078. An oxidoreductase is involved in cercosporin degradation by the bacterium Xanthomonas campestris pv. zinniae. 2006

Taylor, T.V., Mitchell, T.K. and Daub, M.E.

Notes: Fungi of the genus Cercospora are plant pathogens that cause leaf spot and blight diseases, and produce the polyketide toxin cercosporin. The bacterium Xanthomonas campestris is able to rapidly degrade cercosporin. In this study, X. campestris mutants unable to degrade cercosporin were created by chemical mutagenesis. Complementation studies with a plasmid-based library of X. campestris DNA showed that the ability to degrade cercosporin was restored upon transformation with plasmids containing an oxidoreductase gene and a putative transcriptional regulator. These genes were then amplified from the mutant strains by high-fidelity PCR. The PCR products were separated by agarose gel electrophoresis, purified using the Wizard® SV Gel and PCR Clean-Up System, and subcloned into the pGEM®-T Easy Vector. The mutant genes were then sequenced to identify the nature of the mutations. (3531)

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