Total RNA was extracted from the tumors using the Qiagen RNEASY Mini Kit (Qiagen, Germany). Then, 1 μg of the RNA was converted to cDNA, using the First Strand Maxima Synthesis Kit (Thermo Scientific, USA) on a thermal cycler (Labnet, USA). Next, the real-time polymerase chain reaction was conducted using the Power SybrGreen Kit (Invitrogen, USA) on the iCyler IQ5 (Bio-Rad, USA). The sequence of the primers used is detailed in Table 1 . The analysis was done using the iQ5 icycler software, using three housekeeping genes: HPRT, ACTB, and GAPDH.
Icyler iq5
Manufactured by Bio-Rad
Sourced in United States
The ICycler iQ5 is a real-time PCR detection system that enables accurate and sensitive quantification of nucleic acid targets. It features a 96-well format, multiple fluorescence detection channels, and advanced data analysis software. The core function of the ICycler iQ5 is to facilitate real-time PCR experiments and analysis.
Automatically generated - may contain errors
2 protocols using icyler iq5
1
Quantitative Gene Expression Analysis
2
Gene Expression Analysis of Osteogenic Markers
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 at days 7 and 14 and converted to cDNA. Real-time polymerase chain reaction (PCR) was administered using 1X SYBR Premix Ex TaqII on iCyler IQ5 (Bio-Rad, CA, USA), employing a continuous fluorescence-detecting thermal cycler, Rotor Gene Q Sequence Detection System (Qiagen, Hilden, Germany). The primer sequences are listed in Table 1 . Quantification was performed by the standard curve method using REST-RG software, and data were normalized relative to the β-actin housekeeping gene.
List of the Primer Sequences
| Gene | Forward | Reverse |
|---|---|---|
| BMP-2 | TCTGACTGACCGCGTTACTC | TCTCTGTTTCAGGCCGAACA |
| ALP | CAA CAG GGT AGA TTT CTC TTG G | GGT CAG ATC CAG AAT GTT CC |
| OPN | GCC GAG GTG ATA GTG TGG TT | TGA GGT GAT GTC CTC GTC TG |
| β-Actin | ATG CCT GCC GTG TGA AC | ATC TTC AAA CCT CCA TGA TG |
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