Table 1.
The mutation primers used to generate all the mutants
| Mutants | Template | Mutation primers |
|---|---|---|
| M1 (RhoAΔ1-10) |
pECFP-RhoA WT (1-10aa deleted) |
forward: 5’-GTCGACGATTTCGACGTTGGTGATGGAGCC-3’ |
| reverse: 5’-GGCTCCATCACCAACGTCGAAATCGTCGAC-3’, | ||
| M2 (RhoAΔ11-20) |
pECFP-RhoA WT (11-20aa deleted) |
Forward: 5’-CGGAAGAAACTGGTGATTTTGCTCATAGTTAACAGC-3’ |
| Reverse: 5’- GCTGTTAACTATGAGCAAAATCACCAGTTTCTTCCG-3’ | ||
| M3 (RhoAΔ21-30) |
pECFP-RhoA WT (21-30aa deleted) |
Forward: 5’-CTGTGGAAAGACATGCCCAGAGGTGTATGTGC-3’ |
| Reverse: 5’- GCACATACACCTCTGGGCATGTCTTTCCACAG-3’ | ||
| M4 (RhoAΔ31-40) |
pECFP-RhoA WT (31-40aa deleted) |
Forward: 5’- GCGAGGACCAGTTCAACTATGTGGCAG-3’ |
| Reverse: 5’- CTGCCACATAGTTGAACTGGTCCTCGC-3’ | ||
| M5 (RhoAΔ41-50) |
pECFP-RhoA WT (41-50aa deleted) |
Forward: 5’-GCCCACAGTGTTTGAGAAGCAGGTAGAGTTGG-3’ |
| Reverse: 5’- CCAACTCTACCTGCTTCTCAAACACTGTGGGC-3’ | ||
| M6 (RhoAΔ51-60) |
pECFP-RhoA WT (51-60aa deleted) |
Mutant 6 Forward: 5’-CGAGGTGGATGGAGCTGGGCTGGAAG-3’ |
| Mutant 6 Reverse: 5’- CTTCCAGCCCAGCTCCATCCACCTCG -3’ | ||
| M7 (RhoAΔ61-70) |
pECFP-RhoA WT (61-70aa deleted) |
Forward: 5’-CTTTGTGGGACACACCCCTCTCCTACCC-3’ |
| Reverse: 5’-GGGTAGGAGAGGGGTGTGTCCCACAAAG-3’ | ||
| M8 (RhoAΔ71-80) |
pECFP-RhoA WT (71-80aa deleted) |
Forward: 5’-GATTATGATCGCCTGAGGCTGATGTGTTTTTCCATC-3’ |
| Reverse: 5’- GATGGAAAAACACATCAGCCTCAGGCGATCATAATC-3’ | ||
| M9 (RhoAΔ81-90) |
pECFP-RhoA WT (81-90aa deleted) |
Forward: 5’-CCCAGATACCGATGTTATAAGTTTAGAAAACATCCCAG-3’ |
| Reverse: 5’-CTGGGATGTTTTCTAAACTTATAACATCGGTATCTGGG-3’ | ||
| M10 (RhoAΔ91-100) |
pECFP-RhoA WT (91-100aa deleted) |
Forward: 5’-CGACAGCCCTGATCCAGAAGTCAAGC-3’ |
| Reverse: 5’- GCTTGACTTCTGGATCAGGGCTGTCG-3’ | ||
| M11 (RhoAΔ101-110) |
pECFP-RhoA WT (101-110aa deleted) |
Forward: 5’-CAGAAAAGTGGACCCCCATCATCCTGG-3’ |
| Reverse: 5’-CCAGGATGATGGGGGTCCACTTTTCTG-3’ | ||
| M12 (RhoAΔ111-120) |
pECFP-RhoA WT (111-120aa deleted) |
Forward: 5’-CTGTCCCAACGTGCTTCGGAATGATG-3’ |
| Reverse: 5’- CATCATTCCGAAGCACGTTGGGACAG-3’ | ||
| M13 (RhoAΔ121-130) |
pECFP-RhoA WT (121-130aa deleted) |
Forward: 5’-GTTGGGAATAAGAAGGATCTAGCCAAGATGAAGCAG-3’ |
| Reverse: 5’-CTGCTTCATCTTGGCTAGATCCTTCTTATTCCCAAC-3’ | ||
| M14 (RhoAΔ131-140) |
pECFP-RhoA WT (131-140aa deleted) |
Forward: 5’-CACACAAGGCGGGAGCCTGAAGAAGGCAG-3’ |
| Reverse: 5’-CTGCCTTCTTCAGGCTCCCGCCTTGTGTG-3 | ||
| M15 (RhoAΔ141-150) |
pECFP-RhoA WT (141-150aa deleted) |
Forward: 5’-GGAGCCGGTGAAAATTGGCGCTTTTG-3’ |
| Reverse: 5’- CAAAAGCGCCAATTTTCACCGGCTCC-3’ | ||
| M16 (RhoAΔ151-160) |
pECFP-RhoA WT (151-160aa deleted) |
Forward: 5’-GAGATATGGCAAACAGGGCAAAGACCAAAGATGG-3’ |
| Reverse: 5’- CCATCTTTGGTCTTTGCCCTGTTTGCCATATCTC-3’ | ||
| M17 (RhoAΔ161-170) |
pECFP-RhoA WT (161-170aa deleted) |
Forward: 5’-GTGCATGGAGTGTTCATTTGAAATGGCTACG-3’ |
| Reverse: 5’- CGTAGCCATTTCAAATGAACACTCCATGCAC-3’ | ||
| M18 (RhoAΔ171-180) |
pECFP-RhoA WT (171-180aa deleted) |
Forward: 5’-GGAGTGAGAGAGGTTGCTAGACGTGGGAAG-3’ |
| Reverse: 5’- CTTCCCACGTCTAGCAACCTCTCTCACTCC-3’ | ||
| M19 (RhoAΔ181-192) |
pECFP-RhoA WT (181-192aa deleted) |
Forward: 5’-GAGCTGCTCTGCAACTTGTCTTGCCGCG-3’ |
| Reverse: 5’- CGCGGCAAGACAAGTTGCAGAGCAGCTC-3’ |