Total RNA samples were extracted using a RNeasy Midi kit (Qiagen) and aliquot (2 μg) of total RNA was reversed transcribed using a PrimeScriptII 1st strand cDNA Synthesis kit (Takara Bio, Shiga, Japan) according to the standard protocol. Quantitative PCR was performed with RT2 qPCR Primer Assay using the manufacturer’s protocol (Qiagen). Real-Time PCR was carried using SYBR green with the PCR Thermal Cycler MP device (Takara Bio). Each reaction was performed in triplicate. All human qRT-PCR primers were pre-designed, validated RT2 qPCR primer pairs (Qiagen) as follows. For human genes, TYR (PPH01771F), TRP-1 (PPH01770F), dopachrome tautomerase (DCT, PPH16498A), MITF (LPH40195A) and GAPDH (PPH00150F) were used. Relative gene expression was normalized to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA and calculated with the ΔΔCT method.
Rt2 qpcr primer pairs
Manufactured by Qiagen
RT2 qPCR primer pairs are a set of pre-designed and validated primers used for real-time quantitative PCR (qPCR) analysis. They are designed to detect and quantify specific target genes in a sample.
Automatically generated - may contain errors
Lab products found in correlation
Primescript 2 1st strand cdna synthesis kit, by Takara Bio (1 mentions)
Rneasy midi kit, by Qiagen (1 mentions)
Rt2 qpcr primer assay, by Qiagen (1 mentions)
Cfx384 instrument, by Bio-Rad (1 mentions)
Turbo dna free kit, by Thermo Fisher Scientific (1 mentions)
Kapa sybr fast universal master mix, by Roche (1 mentions)
Rneasy mini kit, by Qiagen (1 mentions)
Rt2 ht first strand kit, by Qiagen (1 mentions)
2 protocols using rt2 qpcr primer pairs
1
Quantitative Analysis of Melanogenic Genes
2
Gene Expression Profiling in Infected Cells
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In infected LoVo or JEG-3 cells, RNA was extracted at, respectively, 18 or 20 h p.i. with an RNeasy minikit (Qiagen) as recommended by the manufacturer, using 1 column per well of a 24-well plate and biological duplicates for each condition. Genomic DNA was removed by treatment with a Turbo DNA-free kit (Ambion). cDNAs were generated from 1 to 2 µg total RNA using the RT2 HT first-strand kit (Qiagen/SABiosciences).
qPCR was performed on a CFX384 instrument (Bio-Rad) using Kapa SYBR Fast universal master mix (Kapa Biosystems) as specified by the supplier. Each reaction was performed in triplicate. RT-qPCR primers for YWHAZ, IFI44L, IFITM1, and IL6 were predesigned, validated RT2 qPCR primer pairs from Qiagen/SABiosciences. Other primer pairs were selected from the literature, from primer databases (RTPrimerDB,http://medgen.ugent.be/rtprimerdb/ , or qPrimerDepot, http://primerdepot.nci.nih.gov/ ) and validated for efficiency. Data were analyzed by the ∆∆CT method. Target gene expression data were normalized to the relative expression of the YWHAZ reference gene. The displayed results are representative of three independent experiments in LoVo cells and two experiments in JEG-3 cells.
qPCR was performed on a CFX384 instrument (Bio-Rad) using Kapa SYBR Fast universal master mix (Kapa Biosystems) as specified by the supplier. Each reaction was performed in triplicate. RT-qPCR primers for YWHAZ, IFI44L, IFITM1, and IL6 were predesigned, validated RT2 qPCR primer pairs from Qiagen/SABiosciences. Other primer pairs were selected from the literature, from primer databases (RTPrimerDB,
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