Total and polysomal RNA extracted from NAc were treated with DNase I. Synthesis of cDNA was performed using the AMV reverse transcriptase (Promega) according to the manufacturer’s instructions. The resulting cDNA was used for quantitative real-time PCR. Thermal cycling was performed on an Applied Biosystem 7900HT Fast Real-Time PCR detection system (Applied Biosystems), using a relative calibration curve. The quantity of each mRNA transcript was measured and expressed relative to Glyceraldehyde-3-Phosphate deshydrogenase (GAPDH). The following primers were designed using Primer3 software: Prosapip1: upstream 5′-GTC TGT CAG AAG GAG CAG GC-3′, downstream 5′-CCC CAC GAT CTC ACT CAA CT-3′; Gucy1a3: upstream 5′-CTT CCA CCA AAC TTC CCT A-3′, downstream 5′-GAA CCC ATT ACT TCA ACA CTT A-3′; TAFA-3: upstream 5′-AGA AGG TAA ATC AGC CAT AGT-3′, downstream 5′-ACA GAG GGT GAG CCA AGA-3′; Tsnax: upstream 5′-ATG TGC TCG CTC TAT TGT T-3′, downstream 5′-ATC GGT GAG AAA GGA AAA-3′; GAPDH: upstream 5′-CGA CTT CAA CAG CAA CTC CCA CTC TTC C-3′, downstream 5′-TGG GTG GTC CAG GGT TTC TTA CTC CTT-3′.
Applied biosystem 7900ht fast real time pcr detection system
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Applied Biosystems 7900HT Fast Real-Time PCR detection system is a high-throughput instrument designed for quantitative real-time PCR analysis. It features a 384-well block format and supports rapid thermal cycling to enable fast amplification and detection of nucleic acid targets.
Automatically generated - may contain errors
Lab products found in correlation
Faststart universal sybr green master, by Roche (2 mentions)
Amv reverse transcriptase, by Promega (1 mentions)
Rna extraction kit, by Qiagen (1 mentions)
Superscript 3, by Thermo Fisher Scientific (1 mentions)
Superscript 3 reverse transcriptase, by Thermo Fisher Scientific (1 mentions)
Dnase 1, by Roche (1 mentions)
Allprep dna rna protein kit, by Qiagen (1 mentions)
3 protocols using applied biosystem 7900ht fast real time pcr detection system
1
Quantitative Real-Time PCR Analysis of NAc Gene Expression
2
Quantitative PCR of Transfected N2A Cells
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Total RNA was extracted from transfected N2A cells. RNA extraction was performed using the RNA extraction kit (QIAGEN). Synthesis of cDNA was performed on total RNA with SuperScript III reverse transcriptase (Invitrogen) according to the manufacturer’s instructions. Resulting cDNA was used for quantitative PCR, using Faststart Universal SYBR Green Master (Roche). Thermal cycling was performed on an Applied Biosystem 7900HT Fast Real-Time PCR detection system (Applied Biosystems, Foster city, USA). The primers used are listed in Table S1 .
3
Quantitative RNA Expression Analysis
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Total RNA was extracted from cortices of E14.5 embryos. RNA extraction was performed using the All prep DNA/RNA/protein kit (Qiagen, Hilden, Germany). All RNA samples were treated with DNAse I (Roche, Basel, Switzerland). Synthesis of cDNA was performed on total RNA, which was reverse-transcribed with SuperScript III reverse transcriptase (ThermoFisher Scientific, Waltham, MA, USA) according to the manufacturer’s instructions. Resulting cDNA was used for quantitative PCR, using Faststart Universal SYBR Green Master (Roche). Thermal cycling was performed on an Applied Biosystem 7900HT Fast Real-Time PCR detection system (Applied Biosystems, Foster city, CA, USA). The quantity of each mRNA transcript was measured and expressed relative to Glyceraldehyde-3-Phosphate deshydrogenase (GAPDH). The following primers were designed with Primer3 software: GJA1 forward 5′-GGA CTG CTT TCT CTC ACG TC-3′ and GJA1 reverse 5′-GAG CGA GAG ACA CCA AGG AC-3′; GAPDH forward 5′-GCA CAG TCA AGG CCG AGA AT-3′ and GAPDH reverse 5′-GCC TTC TCC ATG GTG GTG AA-3′.
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