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Abi7900ht pcr machine

Manufactured by Thermo Fisher Scientific
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The ABI7900HT PCR machine is a real-time PCR system designed for accurate and reliable quantification of DNA and RNA targets. It features a 96-well block format and supports a wide range of fluorescent chemistries.

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Lab products found in correlation

Sybr green fluorescent dye, by Thermo Fisher Scientific (2 mentions) Purelink rna columns, by Thermo Fisher Scientific (2 mentions) Neb next ultra rna library prep kit, by Novogene (2 mentions) Trizol, by Thermo Fisher Scientific (2 mentions) Novaseq 6000 pe150, by Novogene (2 mentions) Trizol reagent, by Thermo Fisher Scientific (2 mentions) Superscript 3 first strand synthesis kit for rt pcr, by Thermo Fisher Scientific (2 mentions) Sybr green pcr master mix kit, by Thermo Fisher Scientific (2 mentions) Tri reagent, by Molecular Research Center (2 mentions) Nanodrop 2000c, by Thermo Fisher Scientific (2 mentions) Reliaprep rna cell miniprep system, by Promega (1 mentions) Sybr green, by Thermo Fisher Scientific (1 mentions) Quantitect assays, by Qiagen (1 mentions) Taqman gene expression assay, by Thermo Fisher Scientific (1 mentions) Direct zol rna kit, by Zymo Research (1 mentions) Tgf β1, by Cell Signaling Technology (1 mentions) Hairpin it mirnas qpcr quantitation kit, by GenePharma (1 mentions) Power sybr green pcr master mix, by Takara Bio (1 mentions) Taqman fast universal pcr master mix, by Thermo Fisher Scientific (1 mentions) Taqman primers, by Integrated DNA Technologies (1 mentions)

Close spelling variants of this product

ABI7900HT Fast Real Time PCR machine ABI7900HT machine PCR machine

7 protocols using abi7900ht pcr machine

1

RNA Extraction, Sequencing, and Analysis

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Total RNA was extracted from using TRIzol (Invitrogen) combined with Purelink RNA columns (Fisher) and quantified using a Nano-drop. mRNA was reverse transcribed into cDNA using the ABI High-Capacity cDNA Synthesis kit (ABI). Real-time PCR was performed on an ABI7900HT PCR machine using SYBR green fluorescent dye (Applied Biosystems). Fold changes were calculated using the ΔΔCT method, with Tata Binding Protein (Tbp) mRNA serving as a normalization control. RNaseq libraries were prepared using the NEB Next Ultra RNA Library Prep Kit and sequenced on a NovaSeq 6000 (PE150) (Novogene). RNA-seq reads were aligned to UCSC mm9 genome using STAR aligner (Dobin et al., 2013 (link)) with an option, “–outSAMstrandField intronMotif–outFilterMultimapNmax 1.” Mitochondrial reads were filtered out to avoid sequencing depth bias due to mitochondrial abundance. Then, reads aligned to genes were counted using featureCounts (Liao et al., 2014 (link)). Differential gene expression analysis was performed using edgeR (Robinson et al., 2010 (link)). Hierarchical clustering was performed to identify distinct functional modules of genes using Ward’s criterion and Pearson correlation as a similarity measure. Gene ontology analysis was done using EnrichR (Chen et al., 2013 (link)).
Angueira A.R., Shapira S.N., Ishibashi J., Sampat S., Sostre-Colón J., Emmett M.J., Titchenell P.M., Lazar M.A., Lim H.W, & Seale P. (2020). Early B Cell Factor Activity Controls Developmental and Adaptive Thermogenic Gene Programming in Adipocytes. Cell reports, 30(9), 2869-2878.e4.
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2

RNA Extraction, Sequencing, and Analysis

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Total RNA was extracted from using TRIzol (Invitrogen) combined with Purelink RNA columns (Fisher) and quantified using a Nano-drop. mRNA was reverse transcribed into cDNA using the ABI High-Capacity cDNA Synthesis kit (ABI). Real-time PCR was performed on an ABI7900HT PCR machine using SYBR green fluorescent dye (Applied Biosystems). Fold changes were calculated using the ΔΔCT method, with Tata Binding Protein (Tbp) mRNA serving as a normalization control. RNaseq libraries were prepared using the NEB Next Ultra RNA Library Prep Kit and sequenced on a NovaSeq 6000 (PE150) (Novogene). RNA-seq reads were aligned to UCSC mm9 genome using STAR aligner (Dobin et al., 2013 (link)) with an option, “–outSAMstrandField intronMotif–outFilterMultimapNmax 1.” Mitochondrial reads were filtered out to avoid sequencing depth bias due to mitochondrial abundance. Then, reads aligned to genes were counted using featureCounts (Liao et al., 2014 (link)). Differential gene expression analysis was performed using edgeR (Robinson et al., 2010 (link)). Hierarchical clustering was performed to identify distinct functional modules of genes using Ward’s criterion and Pearson correlation as a similarity measure. Gene ontology analysis was done using EnrichR (Chen et al., 2013 (link)).
Angueira A.R., Shapira S.N., Ishibashi J., Sampat S., Sostre-Colón J., Emmett M.J., Titchenell P.M., Lazar M.A., Lim H.W, & Seale P. (2020). Early B Cell Factor Activity Controls Developmental and Adaptive Thermogenic Gene Programming in Adipocytes. Cell reports, 30(9), 2869-2878.e4.
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3

Quantifying BCR-ABL Fusion Gene Expression

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Total RNA was isolated using the Direct-zol™ RNA Kit (Zymo Research) or ReliaPrep™ RNA Cell Miniprep System (Promega), and synthesis of cDNA was performed (Thermo Fisher). Quantitative real time PCR analyses were performed on ABI7900HT PCR machine (Applied Biosystems) using Quantitect assays (Qiagen) and SYBR Green (Applied Biosystems). The expression of ABL1 and the fusion BCR-ABL (m-bcr; e1-a2) were measured using TaqMan Gene expression assays (Hs01104728_m1 ABL1 and Hs03024844_ft BCR-ABL, respectively) from Applied Biosystems. Relative quantification was calculated using 2−ΔΔCT equation.
Abdelrasoul H., Vadakumchery A., Werner M., Lenk L., Khadour A., Young M., El Ayoubi O., Vogiatzi F., Krämer M., Schmid V., Chen Z., Yousafzai Y., Cario G., Schrappe M., Müschen M., Halsey C., Mulaw M.A., Schewe D.M., Hobeika E., Alsadeq A, & Jumaa H. (2020). Synergism between IL7R and CXCR4 drives BCR-ABL induced transformation in Philadelphia chromosome-positive acute lymphoblastic leukemia. Nature Communications, 11, 3194.
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4

Quantification of miRNA and mRNA

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Total RNA was extracted using the Trizol reagent as recommended by the manufacturer (Invitrogen). RNA quality and concentration were evaluated by spectrophotometry using a NanoDrop 2000c instrument (Thermo Scientific, Rockford, IL, USA). For miRNA analysis, mature miR-141 was detected using a Hairpin-it™ miRNAs qPCR Quantitation kit (GenePharma, Shanghai, China). U6 served as an internal reference. For HIPK2 mRNA analysis, qPCR was performed using the Power SYBR-Green PCR master mix (Takara, Dalian, China) on an ABI 7900HT PCR machine (Applied Biosystems, Foster City, CA, USA), and data were normalized to β-actin and further normalized to the negative control, unless otherwise indicated. Data analysis was performed using the 2−ΔΔCt method. Human recombinant TGF-β1 was purchased from Cell Signaling Technology (no. 8915; Danvers, MA, USA), reconstituted in 20 mM citrate (pH 3.0) at a concentration of 100 μg/ml. Further dilution was made in PBS containing 2 mg/ml albumin, stored at −20°C for future use.
HUANG Y., TONG J., HE F., YU X., FAN L., HU J., TAN J, & CHEN Z. (2014). miR-141 regulates TGF-β1-induced epithelial-mesenchymal transition through repression of HIPK2 expression in renal tubular epithelial cells. International Journal of Molecular Medicine, 35(2), 311-318.
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5

Quantitative Real-Time PCR Protocol

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Total RNA was isolated from tissues or cultured cells using Trireagent (Molecular Research Center, Inc., OH). Reverse transcription was performed using the Invitrogen Superscript III First Strand Synthesis kit for RT-PCR according to the manufacturer’s instructions (Invitrogen, CA), starting with 1μg of total RNA. Primers were added at a final concentration of 80 nM and the sequences are listed in Supplementary Table 1. Quantitative real-time PCR was performed with the SYBR Green PCR master mix kit (Applied Biosystems, CA) using an ABI7900HT PCR machine under “default” conditions: 50°C for 2 minutes and 95°C for 10 minutes, followed by 40 cycles of amplification at 94°C for 15 seconds and 60°C for 1 minute. Melting curves for each primer pair were assessed to ensure specific amplification. The transcript levels were normalized to those of GAPDH.
Huang P., Schulz T.J., Beauvais A., Tseng Y.H, & Gussoni E. (2014). Intramuscular adipogenesis is inhibited by myo-endothelial progenitors with functioning Bmpr1a signaling. Nature communications, 5, 4063.
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6

Quantitative Analysis of miRNA Levels

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Total RNA was extracted from tissue or cells using Trizol reagent (Invitrogen, Thermo Fisher Scientific) followed by reverse transcription for cDNA synthesis from mRNA or miRNA using iScript™ cDNA Synthesis Kit (Bio-Rad, Hercules, CA, USA) and TaqMan® MicroRNA Reverse Transcription Kit (Thermo Fisher Scientific), respectively. The miRNA levels were determined with TaqMan® primers (Integrated DNA Technologies, Coralville, IA, USA) as indicated by quantitative real-time PCR using TaqMan® Fast Universal PCR Master Mix (Thermo Fisher Scientific). All qPCR reactions were run using the Applied Biosystems ABI7900HT PCR machine.
Kuppa S.S., Jia W., Liu S., Nguyen H., Smyth S.S., Mills G.B., Dobbin K.K., Hardman W.J, & Murph M.M. (2018). Autotaxin exacerbates tumor progression by enhancing MEK1 and overriding the function of miR-489-3p. Cancer letters, 432, 84-92.
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7

Quantitative Real-Time PCR Protocol

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Total RNA was isolated from tissues or cultured cells using Trireagent (Molecular Research Center, Inc., OH). Reverse transcription was performed using the Invitrogen Superscript III First Strand Synthesis kit for RT-PCR according to the manufacturer’s instructions (Invitrogen, CA), starting with 1μg of total RNA. Primers were added at a final concentration of 80 nM and the sequences are listed in Supplementary Table 1. Quantitative real-time PCR was performed with the SYBR Green PCR master mix kit (Applied Biosystems, CA) using an ABI7900HT PCR machine under “default” conditions: 50°C for 2 minutes and 95°C for 10 minutes, followed by 40 cycles of amplification at 94°C for 15 seconds and 60°C for 1 minute. Melting curves for each primer pair were assessed to ensure specific amplification. The transcript levels were normalized to those of GAPDH.
Huang P., Schulz T.J., Beauvais A., Tseng Y.H, & Gussoni E. (2014). Intramuscular adipogenesis is inhibited by myo-endothelial progenitors with functioning Bmpr1a signaling. Nature communications, 5, 4063.
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