Talk:20.109(S10):Start-up protein engineering (Day1)

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TR lab

Team colour Mutation X#Z Primer (forward) Silent mutation New restriction site
Teaching example S101L GG AAC GGC TAC ATC CTC GCT GCG CAG TTA CGT CAC G Ala 103 FspI
Teaching experiment M124S GAT GAA GAA GTT GAT GAA AGT ATA CGG GAA GCA GAT ATC GAT GG Arg 126 AccI
Red D20R GAA GCT TTC TCA TTA TTC CGT AAG GAT GGG GAC GGC ACC ATC
[Agi notes: are we ordering RNA or DNA?]
Ala15 HindIII
Orange G25P TTA TTC GAC AAG GAT GGG GAT CCC ACC ATC ACC ACA AAG GAA CTT
[Agi ordered: C GAC AAG GAT GGG GAT CCC ACC ATC ACC ACA AAG GAA C
Primer did not end in G or C, and Tm still is fine.
]
Asp24 BamHI
Yellow Mutation G21W and G23W [Agi notes: actually G23 and G25] primer: C GAC AAG GAT TGG GAC TGG ACC ATC ACC ACA AAG GAA CTT GGC ACC G[Agi ordered: GAC AAG GAT TGG GAC TGG ACC ATC ACC ACA AAG GAA CTT GGT ACC. Primer was too long and missing silent change - I only noticed latter because I had to make sure I wasn't chopping it off.] Silent mutation changes a C to a T at position 302 in the pericam. (position 32 in calmodulin). Enzyme KpnI
Green G23P CA TTA TTC GAC AAG GAT CCG GAC GGC ACC ATC ACC ACA AAG G none BamHI
Blue D129P GTT GAT GAA ATG ATC AGG GAA GCA CCT ATC GAT GGT GAT GGC C Ile 403 BclI
Pink G60P [Agi notes: actually G59P] 5' G CAG GAT ATG ATC AAT GAA GTC GAC GCT GAT CCC AAT GGA ACG 3’ Asp 57 Acc1
Purple N137F 5’ - C GAT GGT GAT GGC CAA GTA TTC TAT GAG GAG TTT GTA CAA ATG – 3' Glu 139 BseRI


WF lab

Team colour Mutation X#Z Primer (forward) Silent mutation New restriction site
Teaching example S101L GG AAC GGC TAC ATC CTC GCT GCG CAG TTA CGT CAC G Ala 103 FspI
Teaching experiment M124S GAT GAA GAA GTT GAT GAA AGT ATA CGG GAA GCA GAT ATC GAT GG Arg 126 AccI
Red E84R G GAC ACA GAC AGC GAG GAG AAA ATC CGA GAA GCA TTC CGT G Glu 82 BseR1
Orange D78K G GCT AGA AAA ATG AAG GAC ACA GAC AGC GAG GAG GAA ATC CG Glu 83 BseRI
Yellow E69K [Agi notes: actually E67K] GGC AAT GGT ACG ATT TAC TTT CCT AAA TTT CTT ACT ATG ATG GC Gly63 RSAI
Green D22H CA TTA TTC GAC AAG CAT GGA GAC GGC ACC ATC Gly 23 BsmBI
Blue Q104R (5’) GC TAC ATC AGC GCT GCT CGG TTA CGA CAC GTC ATG ACA AAC CTC Arg 106 AflIII
Pink E84G 5' G GAC ACA GAC AGC GAG GAG GGA ATC CGA GAA GCA TTC CG Glu 82 BseRI
Purple D56K 5’ CAG GAT ATG ATA AAT GAA GTC AAG GCT GAT GGC AAT GGA ACG 3’ Ile52 BclI


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