Reverse transcription of total RNA was performed using the High Capacity RNA-to-cDNA Master Mix (Applied Biosystems). The quantitative polymerase chain reaction (qPCR) assays for rno-miR-1 and BDNF (assay ID: 02531967_s1) were purchased from Applied Biosystems and applied according to the manufacturer’s protocol. Quantitative polymerase chain reaction conditions were as follows: 50 °C for 2 min, 95 °C for 10 min, 40 cycles of 95 °C for 15 s, 60 °C for 60 s using an ABI 7300HT thermocycler (Applied Biosystems).
Abi 7300ht thermocycler
Manufactured by Thermo Fisher Scientific
Sourced in Germany
The ABI 7300HT thermocycler is a real-time PCR instrument designed for accurate and reliable DNA amplification. It features a Peltier-based thermal block for precise temperature control and a high-performance optical detection system.
Automatically generated - may contain errors
2 protocols using abi 7300ht thermocycler
1
Quantitative Analysis of miR-1 and BDNF Expression
2
Reverse Transcription and qPCR Quantification of miRNAs and mRNAs
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Reverse transcription of total RNA was performed using the High Capacity RNA-to-cDNA Master Mix (Applied Biosystems, Life Technologies, Darmstadt, Germany). The qPCR for miRNA expression was performed with TaqMan®MicroRNA Assays 20X (Applied Biosystems, Life Technologies, Darmstadt, Germany), according to the manufacturer’s protocol: rno-miR-1-3p (assay ID: 002064), U6 (assay ID: 001973), and hsa-miR-21-5p (assay ID: 000397). The qPCR for mRNA expression was performed with TaqMan® Gene Expression Assays 20X (Applied Biosystems, Life Technologies, Darmstadt, Germany), according to the manufacturer’s protocol: HO-1 (Rn01536933_m1), VEGF (Rn01511601_m1), GAPDH (Glyceraldehyde 3-phosphate dehydrogenase) (Rn01462661_g1). QPCR conditions were as follows: 50 °C for 2 min, 95 °C for 10 min, 40 cycles of 95 °C for 15 s, 60 °C for 60 s using an ABI 7300HT thermocycler (Applied Biosystems, Life Technologies, Darmstadt, Germany). The relative expression of miRNAs was calculated using the ΔΔCq-method [18 (link)].
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