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. Author manuscript; available in PMC: 2021 May 19.
Published in final edited form as: Methods Mol Biol. 2019;1982:191–229. doi: 10.1007/978-1-4939-9424-3_12

Table 2.

Primers used in the detection of human NOX isoforms by RT-qPCR

NOX isoform Human sequence of primer Exon location and Tm Integrated DNA technologies Cat. #
NOX1 GGTCAACACGAGGAGAGC
CAAGGATCCACTTCCAAGACTC		 E7F, 60.7
E8R, 61.4		 Hs.PT.49a.435525.4
NOX2 (CYBB) CCCAATCCCTCAGTTTGCT
CCTTCTGTTGAGATCGCCAA		 E7F, 61.1
E8R, 61.2		 Hs.PT.51.3983106
NOX3 ACCTTCTGTAGAGACCGCTAT
TCACATGCATACAAGACCACA		 E7F, 61.2
E8R, 61.1		 Hs.PT.51.3065606
NOX4 CTGTGGTGTTACTATCTGTATTTTCTC
CTTGCTGCATTCAGTTCAACA		 E4F, 60.7
E5R, 60.8		 Hs.PT.51.22695360.g
NOX5 GCCAGTGCCTCAACTTCG
CCACTACCACGTAGCCCATA		 E7F, 62.1
E8R, 62.1		 Hs.PT.58.40874887
DUOX1 GCGTCTACATGAGAAATGCCA
GCAGCAGTGCATCCACAT		 E10F, 61.8
E12R, 61.9		 Hs.PT.51.21346453
DUOX2 CGCCACCTACCAGAACATC
GGTAGAGAAGAACTGC TCAGAG		 E7F, 61.1
E9R, 61.3		 Hs.PT.51.20690545

Primers from PrimeTime™ qPCR Pre-designed Assays (without probes) were purchased from Integrated DNA Technology (IDT) and may have been modified slightly to obtain desired Tm. Primers are listed in the 5′ to 3′ direction (R = reverse and F = forward). The exon number may be different for different accession numbers. The Tm values were determined using IDT’s OligoAnalyzer tool (http://www.idtdna.com/calc/analyzer) using the following final concentrations in the qPCR reaction: [primers] = 200 nM, [Na+] = 55 mM, [Mg2+] = 1.5 mM, and [dNTP] = 0.2 mM. The actual annealing temperatures were determined by performing a gradient RT-qPCR for each primer set using the methods described in the text. These primers detect variant 1 (the full-length variant). To determine whether these primers detect other variants, the user should check the accession number of the variant, some of which are provided in Table 1