Isolation of RNA was performed by using Nucleospin RNA (MACHEREY-NAGEL) following the manufacturer’s protocol. Reverse transcription of mRNA (150 ng total RNA per reaction) was carried out by using TaqMan Reverse Transcription Reagents cDNA kit (Applied Biosystems) according to manufacturer’s protocol. Quantitative analysis of RNA expression by Real Time PCR was performed in the Stratagene MX3005P QPCR System (Agilent Technologies, Germany) by SYBR Green format using the SensiFAST SYBR No-ROX Kit (Bioline) by using 1 µl of cDNA and 0.25 µM primers (see Table S1 ). To exclude non-specificity of the PCR reaction due to contamination or primer dimers, melting curve analysis was performed after each PCR run. Gene expression levels were normalized to Glyceraldehyde 3-phosphate dehydrogenase (GAPDH).
Taqman reverse transcription reagents cdna kit
Manufactured by Thermo Fisher Scientific
Sourced in United States
The TaqMan Reverse Transcription Reagents cDNA kit is a laboratory product designed for the reverse transcription of RNA into complementary DNA (cDNA). It contains the necessary reagents and enzymes to perform this conversion process, which is a fundamental step in various molecular biology and genetic analysis techniques.
Automatically generated - may contain errors
Lab products found in correlation
Nucleospin rna, by Macherey-Nagel (1 mentions)
Sensifast sybr no rox kit, by Meridian Bioscience (1 mentions)
Stratagene mx3005p qpcr system, by Agilent Technologies (1 mentions)
7900 real time pcr system, by Thermo Fisher Scientific (1 mentions)
High capacity cdna rt kit, by Thermo Fisher Scientific (1 mentions)
2 protocols using taqman reverse transcription reagents cdna kit
1
RNA Extraction and qPCR Analysis
2
Quantitative PCR for miR-486-5p and FOXO1
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Isolated RNA was reverse-transcribed to cDNA using TaqMan Reverse Transcription Reagents cDNA kit (Applied Biosystems, USA) and High-capacity RT cDNA kit (Life Technologies, USA) for miR-486-5p and FOXO1, respectively. cDNA was amplified on the 7900 Real-Time PCR System (Applied Biosystems, USA) and the relative expression levels were calculated with the 2 -ΔΔCT method. The PCR conditions were as follows: 95 °C for 10 min followed by 40 cycles at 95 °C for 15 s, and 60 °C for 1 min. Cel-miR-39 (for miR-486-5p) and GAPDH (for FOXO1) were used as an internal reference. The primer sequences for PCR amplification were: miR-486-5p forward 5
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