Table 2.
Primers used in this study.
| Primer name | Sequence 5′- 3′ | Tm* (°C) | Target and notes |
|---|---|---|---|
| Xho_Ptac_mChe.for | AAA ACT CGA GGG GGA ATT CTT GAC AAT TAA TCA TC | 60 | tac promoter; includes a 5′ XhoI restriction site |
| NotI_mChe.rev | AAA GCG GCC GCA AAA ACC GCC CTG CAA GG | 69 | mCherry; includes a 3′ NotI restriction site |
| Ptac_C/YFP.for | AAA ACT CGA GGG GGA ATT CTT GAC AAT TAA TCA TCG GCT CGT ATA ATG TGT GGA ATT GTG AGC GGA TAA CAA TTT TCA CAC AGG AAA CAG CTA AAT GCT GAG CAA GGG CG | 56 | CFP and YFP; the tac promoter is included in the primer |
| Ptac_C/YFP.rev | AAA AGC GGC CGC TAT TAC TTG TAC AGC TCG TCC ATG | 55 | eYFP and eCFP |
| Gib_Ptac+C/YFP.fwd | GTT TTA ATT AAT CAG ATC CCG GGA ATT CTT GAC AAT TAA TC | 48 | tac promoter; contains overlap to SmaI digested pGRG36 |
| Gib_Ptac+C/YFP.rev | GTG GCG GCC GCT ATT GAC CCT TAC TTG TAC AGC TCG TCC | 52 | eCFP and eYFP; contains overlap to SmaI digested pGRG36 |
| Gib_PnptII.fwd | CAC GTT TTA ATT AAT CAG ATC CCA ATT GGG GAT CGG AAG CTT GAC | 58 | nptII promoter from pFru97; contains overlap to SmaI digested pGRG36 |
| Gib_PnptII.rev | CCT TTG CTC ATA TGT TTT TCC TCC TTA TAA AGT TAA TCT TTA GTT AGT TAG GG | 59 | nptII promoter from pFru97; contains overlap to mCherry |
| Gib_mChe.fwd | GGA AAA ACA TAT GAG CAA AGG AGA AGA AGA TAA CAT GG | 58 | mCherry; contains overlap to the nptII promoter |
| Gib_mChe.rev | GGC CGC TAT TGA CCC TTA TTT GTA AAG CTC ATC CA | 47 | mCherry; contains overlap to SmaI digested pGRG36 |
| Gib_PnptII_C/YFP.for | GAA AAA CAT ATG CTG AGC AAG GGC GAG | 62 | eCFP and eYFP; contains overlap to the nptII promoter |
| Gib_PnptII_C/YFP.rev | CTG TAC AAG TAA GGG TCA ATA GCG GCC | 62 | eCFP and eYFP; contains overlap to SmaI digested pGRG36 |
| Gib_C/YFP_PnptII.rev | TGC TCA GCA TAT GTT TTT CCT CCT TAT AAA GTT AAT CTT TAG TTA GTT AGG G | 59 | nptII promoter from pFru97; contains overlap to eCFP and eYFP |
| Gib_PnptII_ngFPs.for | GGC CGC TAT TGA CCC TTA CTT GTA CAG CTC GTC CAT G | 55 | nptII promoter from pRJaph plasmids; contains overlap to SmaI digested pGRG36 |
| Gib_ngFPs.rev | CAC GTT TTA ATT AAT CAG ATC CCC ACG CTG CCG CAA GC | 55 | mTurquoise and eBFP; contains overlap to SmaI digested pGRG36 |
*Tm specific to the overlap to the specified targets, overlaps for isothermal assembly were designed to anneal at 50°C; ABR = antibiotic resistance: Amp = Ampicillin, Kan = Kanamycin, Zeo = Zeocin, Tet = Tetracycline.