Quantitative polymerase chain reaction was performed with the Opticon 2 Real-Time Polymerase Chain Reaction Detection System (Bio-Rad, Hercules, USA) using corresponding primers and the SYBR green Polymerase Chain Reaction Master Mix (Applied Biosystems, Waltham, USA). Reverse-transcribed RNA was amplified by PCR under the following conditions: 95 °C for 5 min, 40 cycles of 30 s at 94 °C, 30 s at 65 °C, 20 s at 72 °C and a final extension step for 10 min at 72 °C. The cycle time values of the genes of interest were first normalized to levels of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in the same sample and gene expression levels were expressed as fold changes versus sham. Primer sequences used are listed in following: MMP-2, 5’-GATCTGCAAGCAAGACATTGTCTT-3’(forward), 5’-GCCAAATAAACCGATCCTTGAA-3’(reverse); MMP-9, 5’-GTAACCCTGGTCACCGGACTT-3’(forward), 5’-ATACGTTCCCGGCTGATCAG-3’ (reverse); GAPDH, 5’-AACCTGCCAAGTATGATGACATCA-3’(forward), 5’-TGTTGAAGTCACAGGAGACAACCT-3’(reverse).
Opticon 2 real time polymerase chain reaction detection system
Manufactured by Bio-Rad
Sourced in United States
The Opticon 2 Real-Time Polymerase Chain Reaction Detection System is a laboratory instrument designed for real-time PCR analysis. It is capable of quantifying and detecting nucleic acid sequences in real-time.
Automatically generated - may contain errors
2 protocols using opticon 2 real time polymerase chain reaction detection system
1
Quantitative PCR Analysis of MMP-2 and MMP-9 Expression
2
Quantifying brain tissue mRNA
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Total mRNA was extracted from the brain tissues around the hematoma by QIAzol Lysis Reagent (miRNeasy Mini Kit; QIAGEN, Gaithersburg, MD) (Li et al., 2017b (link); Su et al., 2017 (link)). One thousand nanograms of RNA from each sample was transcribed into cDNA with the SuperScript VILO cDNA Synthesis Kit (Invitrogen, Carlsbad, United States). Reverse-transcribed RNA was amplified by PCR under the following conditions: 95°C for 5 min, 40 cycles of the 30 s at 94°C, 30 s at 65°C, 20 s at 72°C, and a final extension step for 10 min at 72°C. The quantitative polymerase chain reaction was performed with the Opticon 2 Real-Time Polymerase Chain Reaction Detection System (Bio-Rad, Hercules, United States) using corresponding primers and the SYBR green Polymerase Chain Reaction Master Mix (Applied Biosystems, Waltham, United States). The primer sequences that were utilized are listed in following:
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