Table 1.
Primer sequences for PhIP-seq. Primers for PCRs 1–3 can be used for any PhIP-seq project which is based on the T7Select 10–3b FNS2 vector1, 5. Underlined sequences are adapter sequences, and thus do not participate in the initial round of PCR. The bold sequence is the barcode (“index”) that uniquely defines the sample. We have designed and tested 96 of these sequences; an example (X=1) is shown here. The remaining 95 ad_min_BCX_P7 primer sequences can be found in Supplementary Table 1. The T7-VirScan_SP sequencing primer is specific for analysis of the VirScan library. The Read 2 Illumina Index Read Primer (“Index_SP”) is a standard Illumina primer and available for use from most high throughput DNA sequencing core facilities free of charge.
| Step | Primer Name | Sequence |
|---|---|---|
| PCR1 (Step 42) | T7-Pep2_PCR1_F | 5’-ATA AAG GTG AGG GTA ATG TC-3’ |
| PCR1 (Step 42) | T7-Pep2_PCR1_R+ad_min | 5’-CTG GAG TTC AGA CGT GTG CTC TTC CGA TCA GTT ACT CGA GCT TAT CGT C-3’ |
| PCR2 (Step 45 | T7-Pep2_PCR2_F_P5 | 5’-AAT GAT ACG GCG ACC ACC GAG ATC TAC ACG GAG CTG TCG TAT TCC AGT C-3’ |
| PCR2 (Step 48) | ad_min_BCX_P7 (X is 1 to 96) | 5’-CAA GCA GAA GAC GGC ATA CGA GAT GAC TGA CTG TGA CTG GAG TTC AGA CGT GTG CTC-3’ |
| PCR3 (Step 52) | P5 | 5’-AAT GAT ACG GCG ACC ACC GA-3’ |
| PCR3 (Step 52) | P7.2 | 5’-CAA GCA GAA GAC GGC ATA CGA-3’ |
| Sequencing, Read 1 (Step 55) | T7-VirScan_SP | 5’-GGT GTG ATG CTC GGG GAT CCA GGA ATT CCG CTG CGT-3’ |
| Sequencing, Read 2 (Step 55) | Index_SP | 5’-GAT CGG AAG AGC ACA CGT CTG AAC TCC AGT CAC-3’ |