TABLE 1

PCR primers and conditions for semiquantitative analysis of PXR variants and PXR-induced transcription Nucleotide sequences are reported for primer pairs used in RT-PCR analysis of PXR transfection experiments in HepG2 and Caco-2 cells. Two common primers are used for the UGT2B isoform PCR: cR1 has two mismatches to UGT2B4 and one mismatch each to UGT2B7, UGT2B11, and UGT2B15. cR2 has one mismatch to UGT2B10 and three mismatches to UGT2B28. The nucleotide sequences for UGT1A4F, UGT1A6F and R, UGT1A10R, and all UGT2B primers except UGT2B28F were obtained from Congiu et al. (2002). The sense primer for UGT1A10 was designed by Strassburg et al (1997). PXR variant primers were designed by J.-M. Heydel.

Target Transcript
Primer Set
Annealing Temp.
Amplicon Size
No. of Cycles on HepG2 cDNA
No. of Cycles on Caco-2 cDNA
°C bp
T1,T2,T3 FC: 5′ agaaggagatgatcatgtccga 3′ 60 359 32 34
RC: 5′ gtttgtagttccagacactgcc 3′ 248
T1 F1: 5′ caagccaagtgttcacagtgag 3′ 60 818 35 35
R1: 5′ caaagagcacagatcttccg 3′
T2 F2: 5′ gcagcatgacagtcacc 3′ 62 460 35 nd
R2: 5′ ctccttcttcatgccgctct 3′
T3 F3: 5′ cactgcctttacttcagtggg 3′ 60 764 35 nd
R3: 5′ cagctgcagagagacccg 3′
PXR F: tgtcatgacatgtgaaggatg 58 327 25 25
R: ttgaaatgggagaaggtagtg
CYP3A7 F: agttgctatgagacttgagag 50 637 30 32
R: aatctacttccccagcactga
UGT1A F: tgaaagcatatgcaatggcgt 50 466 30 27
R: tcaatgggtcttggatttgtg
UGT1A1 F: atgctgtggagtcccagggc 50 932 30 30
R: ccattgatcccaaagagaaaacc
UGT1A3 F: atggcaatgttgaacaatatg 58 247 35 35
R: ggtctgaattggttgttagtaatc
UGT1A4 F: acgctgggctacactcaagg 66 200 40 35
R: gacaggtacttagccagcacc
UGT1A6 F: cttttcacagacccagccttac 58 289 42 25
R: tatccacatctctcttgaggacag
UGT1A7 F: tggctcgtgcagggtggactg 63 310 nd 35
R: ttcgcaatggtgccgtccagc
UGT1A8 F: ctgctgacctgtggctttgct 63 248 nd 35
R: ccattgagcatcggcgaaat
UGT1A9 F: gaggaacatttattatgccaccg 50 281 34 32
R: ccattgatcccaaagagaaaacc
UGT1A10 F: cctctttcctatgtccccaatga 63 205 nd 35
R: gcaacaaccaaattgatgtgtg
UGT2B F: aagttctaggaagacccactac 58 205 30 na
R: caccacaacaccattttctcca
UGT2B4 F: tctactcttaaatttgaagtttatcctgt 58 278 30 na
cR1: tcagcccagcagctcaccacaggg
UGT2B7 F: agttggagaatttcatcatgcaacaga 58 232 26 30
cR1: tcagcccagcagctcaccacaggg
UGT2B10 F: tgacatcgtttttgcagatgctta 58 152 28 na
cR2: caggtacataggaaggagggaa
UGT2B11 F: cttccattctttttgatcccaatgatg 58 307 30 na
cR1: tcagcccagcagctcaccacaggg
UGT2B15 F: gtgttgggaatattatgactacagtaac 58 141 32 na
cR1: tcagcccagcagctcaccacaggg
UGT2B17 F: gtgttgggaatattctgactataatata 58 242 41 na
cR2: caggtacataggaaggagggaa
UGT2B28 F: atcccaatgacgcattcactcttaaactc 58 340 nd na
cR2: caggtacataggaaggagggaa
β-actin1 F: ctggcggcaccaccatgtaccct 50 205 18 18
R: ggaggggccggactcgtcatact
β-actin2 F: cgtaccactggcatcgtgat 58 452 18 18
R: gtgttggcgtacaggtcttt
GAPDH F: 5′-acccactcctccacctttg-3′ 64 178 25 25
R: 5′-ctcttgtgctcttgctggg-3′

  • F, forward primer; R, reverse primer; bp, base pair; nd, not detected; na, not attempted.