Then, q-PCR was carried out using SYBR Master Mixture (TaKaRa, Ohtsu, Japan) and TaKaRa Thermal Cycler Dice Real-Time PCR System TP800. The primer sequences were as follows: 5′-GTGGTGGAGATGACTGAAGCC-3′ (sense) and 5′-CCACGGGGACTGTTATTCG-3′ (antisense) were for ANXA2 (110 bp). Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used as an internal control; 5′-TGACTTCAACAGCGACACCCA-3′ (sense) and 5′-CACCCTGTTGCTGTAGCCAAA-3′ (antisense) were for GAPDH (121 bp). The thermal cycling conditions consisted of 1 cycle at 95°C for 30 s, followed by 45 cycles at 95°C for 5 s and 60°C for 30 s. Data were analyzed by the comparative threshold (2-∆∆Ct) method.
Thermal cycler dice real time pcr system tp800
Manufactured by Takara Bio
Sourced in Japan
The Thermal Cycler Dice Real-Time PCR System TP800 is a laboratory instrument designed for real-time polymerase chain reaction (PCR) analysis. It features a compact design and provides precise temperature control for accurate DNA amplification and quantification.
Automatically generated - may contain errors
Lab products found in correlation
Isogen, by Nippon Gene (2 mentions)
Sybr master mixture, by Takara Bio (1 mentions)
Taqman microrna reverse transcription kit, by Thermo Fisher Scientific (1 mentions)
Taqman microrna assay, by Thermo Fisher Scientific (1 mentions)
Primescript 1st strand cdna synthesis kit, by Takara Bio (1 mentions)
Tb green premix ex taq 2, by Takara Bio (1 mentions)
4 protocols using thermal cycler dice real time pcr system tp800
1
Quantitative PCR Analysis of ANXA2
2
qRT-PCR Analysis of NPC1L1 and ACTB
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qRT-PCR was conducted using SYBR Green EX (Takara Bio, Shiga, Japan) and the Thermal Cycler DICE Real-Time PCR System TP800 (Takara). The primers used for the analysis of each of the mRNAs were as follows: NPC1L1 (forward, 5′-GACCGGCCCAACATCAA-3′ , reverse, 5′-CCGCAGAGCTTCTGTGTAATC-3′ ), ACTB (forward, 5′-GCGTGACATTAAGGAGAAG-3′ , reverse, 5′-GAAGGAAGGCTGGAAGAG-3′ ). The amount of each mRNA was normalized relative to that of ACTB.
3
Quantifying miR-142-5p Expression in Mouse Bone Marrow
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Total RNA was extracted from mouse bone marrow (BM) samples using ISOGEN (Nippon Gene). cDNA synthesis was performed from 10 ng of total RNA using a TaqMan MicroRNA Reverse Transcription kit (Applied Biosystems), according to the manufacturer's instructions. Thereafter, miRNA expression levels were quantified using a TaqMan MicroRNA Assay (Applied Biosystems) for miR‐142‐5p in a Thermal Cycler Dice TP800 Real‐Time PCR system (TaKaRa). Each assay was performed in technical triplicates. miR‐142 expression levels were normalized to Actb/β‐actin as an internal control. The average threshold cycle (Ct) for three replicates per sample was used to calculate ΔCt. Finally, the relative quantification for gene expression was calculated using the 2−ΔΔCt method.
4
RNA Extraction and qRT-PCR Analysis
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Total RNA was extracted from mouse BM samples and hematopoietic cells using ISOGEN (Nippon Gene), and 500 ng of RNA was converted to cDNA using a PrimeScript 1st strand cDNA Synthesis Kit (6110A; TaKaRa). cDNA aliquots were used for quantitative RT‐PCR (RT‐qPCR). For RT‐qPCR, TB Green Premix Ex Taq II (RR820A; TaKaRa) was used as the Taq polymerase, and reactions were performed using the intercalator method in a Thermal Cycler Dice TP800 Real‐Time PCR system (TaKaRa). In experiments using relative quantification, expression levels were normalized to Actb/β‐actin expression. The sequences of all primers used in this study are listed in Table S2 .
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