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Realmastermix

Manufactured by Eppendorf
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Sourced in Germany, United States

RealMasterMix is a premixed, ready-to-use solution for real-time PCR (polymerase chain reaction) applications. It contains all the necessary components, including a DNA polymerase, buffer, and fluorescent dye, to perform accurate and efficient real-time PCR experiments.

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Lab products found in correlation

Iscript cdna synthesis kit, by Bio-Rad (4 mentions) Mastercycler realplex2, by Eppendorf (3 mentions) Genorm kit, by Primerdesign (1 mentions) Illustra tripleprep kit, by GE Healthcare (1 mentions) Rneasy mini kit, by Qiagen (1 mentions) Turbo dnase 1, by Thermo Fisher Scientific (1 mentions) Rneasy micro kit, by Qiagen (1 mentions)

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SYBR Green RealMasterMix (10 citations)

6 protocols using realmastermix

1

Quantifying ALK mRNA Expression

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Expression of ALK mRNA was analyzed in both NB cell lines and samples by using 25 ng of cDNA in two independent experiments, in duplicate, on EpGradient Realplex PCR System (Eppendorf), using RealMasterMix (Eppendorf). We used specific 6FAM labeled TaqMan Assays (Applied Biosystems) for ALK, and employed the geNORM Kit (Primerdesign) to choose the best reference genes (18S, GAPDH and UBC). The comparative ΔCt method was then applied, normalizing ALK threshold cycle values on the geometric mean of the three housekeeping genes. All qPCR analyses were conducted in accordance to the MIQE guidelines for qPCR research reporting [68 (link)], as detailed in Supplementary Table 4.
De Mariano M., Stigliani S., Moretti S., Parodi F., Croce M., Bernardi C., Pagano A., Tonini G.P., Ferrini S, & Longo L. (2017). A genome-wide microRNA profiling indicates miR-424-5p and miR-503-5p as regulators of ALK expression in neuroblastoma. Oncotarget, 8(34), 56518-56532.
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2

Quantifying Gene Expression in Mouse VTA

Cited 4 times
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Mice were sacrificed either 24 h or 2 months after the last injection of VEH, MPH, FLX, MPH+FLX, or cocaine. Brains were extracted and sliced into 1-mm diameter coronal sections at which time an 18-gauge needle was used to dissect out bilateral VTA punches that were then stored at −80 °C until use. RNA was isolated using Illustra TriplePrep kit (GE Healthcare) according to the manufacturer’s instructions (Valladares et al. 2010 (link)). cDNA was then created from these samples using iScript cDNA synthesis kit (Bio-Rad). Quantitative real-time PCRs were performed in duplicates using 96 well PCR plates and RealMasterMix (Eppendorf) with an Eppendorf MasterCycler Realplex2, according to the manufacturer’s instructions. Threshold cycle [C(t)] values were measured using the supplied software and analyzed with the ΔΔC(t) method as previously described (LaPlant et al. 2010 (link); Vialou et al. 2010 (link); Warren et al. 2013 (link)). Primer sequences for ERK2 (Mapk1), BDNF (Bdnf), CREB (Creb1), Zif268 (Egr1), cFos (Fos), mTOR (Mtor), and glyceraldehyde-3-phosphate dehydrogenase (Gapdh) are listed in Table 1.
Alcantara L.F., Warren B.L., Parise E.M., Iñiguez S.D, & Bolaños-Guzmán C.A. (2014). Effects of psychotropic drugs on second messenger signaling and preference for nicotine in juvenile male mice. Psychopharmacology, 231(8), 1479-1492.
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3

Quantifying Cardiac Gene Expression in Mice

Cited 2 times
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Total mRNA from mouse hearts and aortas was isolated by the TriZol method [35 (link), 37 ]. One µg total mRNA was reverse transcribed into cDNA and analyzed by real-time PCR using RealMasterMix and realplex2 Mastercycler (Eppendorf AG, Hamburg, Germany) [38 (link)]. To analyze the expression of interleukin (IL)-1β, CXCL1, CXCL2, transforming growth factor beta (TGFβ) 1, TGFβ3, connective tissue growth factor (CTGF), collagen (Coll) IaI, Coll Ia2 and Coll 3 mRNA we used predesigned probe sets (Integrated DNA Technologies, Coralville, IA). Variations in loading were adjusted using hypoxanthine-guanine phosphoribosyltransferase (HPRT) mRNA expression.
Antoniak S., Cardenas J.C., Buczek L.J., Church F.C., Mackman N, & Pawlinski R. (2016). Protease-activated receptor 1 contributes to angiotensin II-induced cardiovascular remodeling and inflammation. Cardiology, 136(4), 258-268.
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4

Quantification of Transcriptional Factors in NAc

Cited 2 times
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Mice were sacrificed either 24-h after the last exposure to CON, ES, or PS and 1.25 mm punches were taken from NAc (including both core and shell) and stored at − 80°C until use. RNA was isolated using RNEasy Micro kits (Qaigen) and cDNA was created from these samples using iScript cDNA synthesis kit (Bio-Rad; Hercules, California). qPCRs were performed in triplicate using 96 well PCR plates and RealMasterMix (Eppendorf) with an Eppendorf MasterCycler Realplex2 according to manufacturer’s instructions. Threshold cycle [C(t)] values were measured using the supplied software and analyzed with the ΔΔC(t) method as described previously [27 (link),46 (link)]. Primer sequences for Mapk1 (ERK2), Deltafosb (ΔFosB), Creb1 (CREB), and Gapdh were generated from the Harvard Primer Bank and are listed in Table 1.
Warren B.L., Sial O.K., Alcantara L.F., Greenwood M.A., Brewer J.S., Rozofsky J.P., Parise E.M, & Bolaños-Guzmán C.A. (2014). Altered gene expression and spine density in nucleus accumbens of adolescent and adult male mice exposed to emotional and physical stress. Developmental neuroscience, 36(0), 250-260.
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5

Quantifying mRNA Levels of rpoN and hrpL

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The mRNA levels of rpoN and hrpL were measured by qRTPCR. Briefly, bacterial cells cultured in MM for 12 h were harvested and total RNA was isolated using RNeasy mini kit (Qiagen, Hilden, Germany) according to the manufacturer’s instruction. Extracted RNA was treated with Turbo DNase I (Ambion, Austin, TX), and cDNA was synthesized using iScript cDNA synthesis kit (Bio-Rad Laboratories, Hercules, CA). The complementary (c)DNA level of target genes was quantified by qRT-PCR using a Real Master Mix (Eppendorf, Westbury, NY, USA), as described previously (Peng et al., 2006 (link)). Data were analysed using A RELATIVE EXPRESSION SOFTWARE TOOL (Pfaffl et al., 2002 (link)). The expression level of rplU was used as an endogenous control for data analysis (Mah et al., 2003 (link)).
Yuan X., Khokhani D., Wu X., Yang F., Biener G., Koestler B.J., Raicu V., He C., Waters C.M., Sundin G.W., Tian F, & Yang C.H. (2015). Cross-talk between a regulatory small RNA, cyclic-di-GMP signalling and flagellar regulator FlhDC for virulence and bacterial behaviours. Environmental microbiology, 17(11), 4745-4763.
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6

Hippocampal Gene Expression Analysis

Cited 2 times
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Whole hippocampus was microdissected on dry ice and stored at −80°C until assayed [45 (link)]. RNA isolation was carried out with RNEasy Micro kits according to the manufacturer’s instructions (Qiagen; Austin, TX). RNA was then reverse transcribed into cDNA, using the iScript cDNA synthesis kit (Bio-Rad; Hercules, CA). qPCR was then conducted using a commercially available kit (RealMasterMix, Eppendorf; Westbury, NY), running duplicate samples from each animal. Cycle threshold (Ct) values were determined and changes in gene expression were analyzed by the ΔΔCt method [46 (link)], using glyceraldehyde 3-phosphate dehydrogenase (GAPDH) as housekeeping reference. The primer sequences for the analyzed genes are displayed in Table 1.
Iñiguez S.D., Flores-Ramirez F.J., Themann A, & Lira O. (2020). Adolescent fluoxetine exposure induces persistent gene expression changes in the hippocampus of adult male C57BL/6 mice. Molecular neurobiology, 58(4), 1683-1694.
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