Total RNA was extracted from cells using TRIzol® reagent (Invitrogen; Thermo Fisher Scientific, Inc.), and cDNA samples were synthesized using random primers and a Reverse Transcriptase PCR kit (Takara Bio, Inc.) according to the manufacturer's protocols. cDNA was used as a template for semi-quantitative PCR. The thermocycling conditions were as follows: 94°C for 5 min, 94°C for 30 sec, 68°C for 30 sec and 72°C for 12 cycles with a decrease in 1°C/cycle; then, 94°C for 30 sec, 55°C for 30 sec, and 72°C for 30 sec for 18-27 cycles depending on the abundance of the target genes. The PCR products were identified by electrophoresis using 1.5% agarose gels. GAPDH was used as an internal reference control. The results were recorded using Gel Doc 1000 imaging system and Quantity One version 4.5.0 software (Bio-Rad Laboratories, Inc.). The primers used in this study are shown in Table I .
Gel doc 1000 imaging system
Manufactured by Bio-Rad
Sourced in United States
The Gel Doc 1000 imaging system is a compact and versatile device designed for capturing and analyzing gel images. It offers automated image acquisition and analysis capabilities, allowing users to visualize and document their gel-based experiments, such as DNA, RNA, or protein electrophoresis.
Automatically generated - may contain errors
Lab products found in correlation
Trizol reagent, by Thermo Fisher Scientific (2 mentions)
Reverse transcriptase pcr kit, by Takara Bio (1 mentions)
Taqman reverse transcription kit, by Takara Bio (1 mentions)
T100 thermal cycler, by Bio-Rad (1 mentions)
Mygenie 96 thermal block, by Bioneer (1 mentions)
Accupower hotstart pcr premix kit, by Bioneer (1 mentions)
3 protocols using gel doc 1000 imaging system
1
Semi-Quantitative PCR for Gene Expression
2
Quantification of S100A4 Expression in CRC
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Total RNA from human CRC tissues was isolated using Trizol Reagent (Invitrogen, CA, USA). RT-PCR was performed by using the TaqMan reverse transcription kit according to the manufacturer's instructions (TaKaRa Biotechnology, Dalian, China) and the T100 Thermal Cycler (Bio-Rad, CA, USA). PCR products were identified by electrophoresis with 1.5% agarose gels and recorded using the Gel Doc 1000 imaging system (Bio-Rad, CA, USA). GAPDH RNA was served as an input control. The following primers were used: S100A4, forward primer: 5'-TCAGAACTAAAGGAGCTGCTGACC-3', reverse primer: 5'-TTTCTTCCTGGGCTGCTTATCTGG-3'; GAPDH, forward primer: 5'-CAGCGACACCCACTCCTC-3', reverse primer: 5'-TGAGGTCCACCACCCTGT-3'.
3
Quantitative RT-PCR Analysis of Gene Expression
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Total RNA was extracted with TRIzol reagent according to the manufacturer’s instructions. Total RNA (1 µg) in the final 20 µL cDNA system was synthesized by reverse transcription synthesis kit (#04897030001; Roche Diagnostics, Mannheim, Germany). A semi-quantitative reverse transcriptase–polymerase chain reaction (sqRT–PCR) was performed using an AccuPower HotStart PCR PreMix kit according to the manufacturer’s instructions (K-5051; Bioneer, Daejeon, Korea) in a MyGenie 96 Thermal Block (Bioneer, Daejeon, Korea). The cycle sequence of each PCR reaction was performed using an initial denaturation at 94 °C for 5 min, followed by 28 cycles at 94 °C for 30 sec, 52 °C for 30 sec, and 72 °C for 30 sec. After 28 cycles, an additional elongation step was performed at 72 °C for 7 min. PCR products were identified by electrophoresis with 1.0% agarose gels and recorded using a Gel Doc 1000 imaging system (Bio-Rad, Carlsbad, CA, USA). GAPDH RNA served as a housekeeping gene relative to the control.
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