Two-Hybrid Primers
Primers used for amplifying yeast ORFs and their "common tails":
Forward primer:
5' AA TTC CAG CTG ACC ACC ATG xxx xxx xxx xxx xxx xxx xxx 3'
whereas the ATG
represents the actual start codon of the yeast ORF.
Reverse primer:
5' xxx xxx xxx xxx xxx xxx *** CAT
GGC AAT TCC CGG GGA TC 3' (coding strand)
3' xxx xxx xxx xxx xxx xxx *** GTA
CCG TTA AGG GCC CCT AG 5' (actual sequence of primer)
whereas the xxx's represent the ORF-specific
sequences and the ***'s one of the 3 stop codons.
For the re-PCRs the following primers have been used:
Forward primer:
5' C TAT CTA TTC GAT GAT GAA GAT ACC CCA
CCA AAC CCA AAA AAA GAG ATC GAA
TTC CAG CTG ACC ACC ATG 3'
Reverse primer:
5' CATGGCAATTCCCGGGGATCCGTCGACCTGCAGAGATCTATGAATCGTAGATACTGAAAAACCCCGCAAG 3' (coding strand)
3' GTACCGTTAAGGGCCCCTAGGCAGCTGGACGTCTCTAGATACTTAGCATCTATGACTTTTTGGGGCGTTC 5' (actual primer)