RAW264.7 cells were seeded at 1×105 cells in 6-well plates, and total RNA was extracted and purified on day 4 by an RNA purification kit according to the manufacturer’s protocol (Favorgen Biotech Corp., Ping Tung, Taiwan). The complementary DNA was synthesized from 0.5 mg of total RNA by the iScript cDNA synthesis kit (Bio-rad, Hercules, CA, USA). The primer sequences of cathepsin K, DC-STAMP, MMP-9, and GAPDH are shown in Table 1 . Quantitative real-time PCR was measured using KAPA SYBR fast (KAPA biosystem, Wilmington, MA, USA) on Bio-rad real-time PCR CFX96 (Bio-rad). The gene expression values were calculated from the differences between Ct of the target’s gene with GAPDH and expressed as relative fold change (the 2-ΔΔCT method). The value of undifferentiated cells was defined as 1.
Rna purification kit
Manufactured by Favorgen Biotech
The RNA purification kit is a laboratory tool designed to extract and purify ribonucleic acid (RNA) from various biological samples. It employs a rapid and efficient method to isolate high-quality RNA suitable for downstream applications such as gene expression analysis, reverse transcription, and other molecular biology techniques.
Automatically generated - may contain errors
Lab products found in correlation
Kapa sybr fast, by Roche (1 mentions)
Iscript cdna synthesis kit, by Bio-Rad (1 mentions)
Mic qpcr cycler, by Bio Molecular Systems (1 mentions)
Dnase free rnase, by Merck Group (1 mentions)
First strand cdna synthesis kit, by Thermo Fisher Scientific (1 mentions)
Luna universal qpcr master mix, by New England Biolabs (1 mentions)
2 protocols using rna purification kit
1
Quantifying Osteoclastogenesis Gene Expression
2
Quantitative RT-PCR Gene Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Total RNA was extracted from cultured cells using an RNA Purification Kit (Favorgen, Ping-Tung, Taiwan), and 200 ng of total RNA was treated with RNase-free DNase (Sigma Aldrich, St. Louis, MO, USA) for 15 min. After inactivation of DNase with EDTA treatment and heating, total RNA was reverse transcribed into cDNA using First Strand cDNA Synthesis Kit (Thermo Fisher Scientific, Waltham, MA, USA). Quantitative RT-PCR was performed on cDNA samples with Luna® universal qPCR master mix (New England Biolabs, Ipswich, MA, USA) on the Mic qPCR Cycler (bio molecular systems, Queensland, Australia). The relative mRNA level was quantitated as values of 2(Ct[RPL32] − Ct[gene of interest]). The sequences of the forward and reverse primers are listed in Table 1 .
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