TABLE 1.
Sequences of gene specific primers and hybridization probes used in quantitative PCR analysis
| Target mRNA | Assay Format | Primer/Probe | Sequences (5′ to 3′) |
|---|---|---|---|
| ErbB1 (EGFR) | SYBR Green I | Forward primer (PC4886) | CCACCTGTGCCATCCAAACT * |
| Reverse primer (PC4887) | GGCGATGGACGGGATCTT * | ||
| ErbB2 | SYBR Green I | Forward primer (PC4888) | AGCCTTGCCCCATCAACTG * |
| Reverse primer (PC4889) | AATGCCAACCACCGCAGA * | ||
| ErbB3 | SYBR Green I | Forward primer (PC4890) | CCCTGCCATGAGAACTGCAC * |
| Reverse primer (PC4891) | TCACTGTCAAAGCCATTGTCAGAT * | ||
| ErbB4 | SYBR Green I | Forward primer in TK domain (PC1542) | CCTGGAAGAAAGACGACTCGTTC |
| Reverse primer in TK domain (PC1541) | CGTCACTCTGATGGGTGAATTTCC | ||
| ErbB4 JM region | Hybridization Probe | Forward primer flanking JM region (PC2338) | CAGTGTGAGAAGATGGAAGATG |
| Reverse primer flanking JM region (PC5221) | CCACAATGACCAGAATGAAGAG | ||
| JM-a | Hybridization Probe | Donor-probe F1 (PC5213) | GCCATCCAAACTGCACCCAA-Flu |
| Acceptor-probe R1 (PC5215) | Red610-GTAACGGTCCCACTAGTCATGACTGCATTTA-p | ||
| JM-b | Hybridization Probe | Donor-probe F2 (PC5214) | AAGACTGCATCGGCCTGATGGATA-Flu |
| Acceptor-probe R2 (PC5216) | Red640-AACTCCCCTGATTGCAGCTGGA-p | ||
| JM-c | Hybridization Probe | Donor-probe F1 (PC5213) | GCCATCCAAACTGCACCCAA-Flu |
| Acceptor-probe R2 (PC5216) | Red640-AACTCCCCTGATTGCAGCTGGA-p | ||
| JM-d | Hybridization Probe | Donor-probe F2 (PC5214) | AAGACTGCATCGGCCTGATGGATA-Flu |
| Acceptor-probe R1 (PC5215) | Red610-GTAACGGTCCCACTAGTCATGACTGCATTTA-p | ||
| ErbB4 CYT region | Hybridization Probe | Forward primer flanking CYT region (PC5268) | CCAGTCCAAATGACAGCAAG |
| Reverse primer flanking CYT region (PC5269) | CCTTGTTCAGCAGCAAAACC | ||
| CYT-1 | Hybridization Probe | Donor-probe F3 (PC5217) | CTTCCAGAGCAAGAATTGACTCGAATAGG-Flu |
| Acceptor-probe R3 (PC5218) | Red610-GAAATTGGACACAGCCCTCCTCC-p | ||
| CYT-2 | Hybridization Probe | Donor-probe F3 (PC5217) | CTTCCAGAGCAAGAATTGACTCGAATAGG-Flu |
| Acceptor-probe R4 (PC5219) | Red640-CCAGTTTGTATACCGAGATGGAGGTTTTG-p | ||
| TACE | SYBR Green I | Forward primer (PC5128) | GGATGTGAAGATGTTGCTAGAG |
| Reverse primer (PC5129) | CTGGGCTATAATAAGCCTTTG | ||
| Presenilin-1 | SYBR Green I | Forward primer (PC5140) | GCCCTGCACTCAATTCTG |
| Reverse primer (PC5141) | ACACTTCCCCCAAGTAAATG | ||
| Presenilin-2 | SYBR Green I | Forward primer (PC5142) | TCCTCTTCACCTATATCTACCTTG |
| Reverse primer (PC5144) | CACAGCCACGAGATCATAC | ||
| Nicastrin | SYBR Green I | Forward primer (PC5134) | AGCCTCTCCCACCATCTTC |
| Reverse primer (PC5135) | CATACAGAGCACGTCCCAG | ||
| Pen-2 | SYBR Green I | Forward primer (PC5145) | TTCCTTGTCCCAGCCTAC |
| Reverse primer (PC5146) | GGTGAGCACTATCACCCAG | ||
| Aph-1a | SYBR Green I | Forward primer (PC5151) | GGTTAGCATCGCTGAGTGAG |
| Reverse primer (PC5152) | GAAAGGCTGAAGTCAGGAAG | ||
| H6PD | SYBR Green I | Forward primer (PC5242) | GATCCTGCCTTTCCGAGAC |
| Reverse primer (PC5244) | CCGCACTGCTGACATTG | ||
| 18S rRNA | SYBR Green I | Forward primer (PC1745) | GTGGAGCGATTTGTCTGGTT |
| Reverse primer (PC1746) | CGCTGAGCCAGTCAGTGTAG |
The donor probes were labelled with Fluorescein (Flu) dye at the 3′end. The acceptor probes were labelled with LightCycler Red610 (R610) or Red640 (R640) at the 5′ end, and the 3′ hydroxyl group of the probe was blocked with a phosphate (p). TACE: tumor necrosis factor-α-converting enzyme; H6PD: hexose 6-phosphate dehydrogenase.
*
Primers indicated are as designed by Junttila et al (36).