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Table 1 The mutation primers used to generate all the mutants

From: Enzymatically active Rho and Rac small-GTPases are involved in the establishment of the vacuolar membrane after Toxoplasma gondii invasion of host cells

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’