User:Jerome Bonnet/Notebook/Part Collection
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From the genetic memory project, this a repository of the parts I will depose in the registry of standard biological parts. They will serve to test for the activity and crosstalk of the different Integrases and Excisionases .
Sequences are displayed in the Registry format, meaning that Biobrick prefix and suffix are omitted.
Note: ATG and Stop removed from protein sequence, encoded in Freiburg prefix and suffix. Freiburg standard Add AG amino acids in N-ter after the Methionine and TG before the stop. After checking on litterature and with Alfonso that should not be a problem ( functional N or C terminal fusions of Integrases have been made).
Bbw_1 Bxb1 integrase
This part encodes an integrase from the Serine Recombinase family. Bxb1 integrase catalyse integration of a given sequence by recombination between specific recombination sites attB and attP. In association with a specific subunit, called excisionase
Bbw_2 Bxb1 excisionase >>>> Received
Bbw_3 TP901-1 integrase >>>>> Received
Bbw_4 TP901-1 excisionase >>>>> Received
Bbw_5 PhiRV1 integrase
Bbw_6 PhiRV1 excisionase
Note: RBS for integrase is B0030 (efficiency 0,6). Excisionases are controlled by RBS B0034 (efficiency 1). this should ensure expression of the excisionase over the stochiometric ratio with integrase, and force the excision-like reaction. Hope it will be enough, but I wanted also to be sure of a correct integrase expression for the first experiment...
Bbw_7 Bxb1 generator: RBS(B0030)-Bxb1 integrase-6His
Bbw_8 Bxb1 Int/Xis generator: RBS(B0030)-Bxb1 integrase-6His / RBS B0034-Bxb1 excisionase-6his
Bbw_9 TP901 generator: RBS(B0030)-TP901-1 integrase-6His
Bbw_10 TP901 Int/Xis generator: RBS(B0030)-TP901 integrase-6His / RBS B0034-TP901 excisionase-6his
Bbw_11 PhiRV1 generator: RBS(B0030)-PhiRV1 integrase-6His
Bbw_12 PhiRV1 Int/Xis generator: RBS(B0030)-PhiRV1 integrase-6His / RBS B0034-PhiRV1 excisionase-6his
Note: a random DNA sequence checked for: no ATG, no RBS like sequence, no TATAA box, no terminators no apparent strange RNA structure...and blasted to verify no relevant homology (pretty unique). Bear an inverted constitutive promoter J23119, from the constitutive promoters family, the stronger one. flanked by att sites for each recombinases, BP and LR.
Bbw_13 Bxb1-BP flippee >>>>> Received
Bbw_14 Bxb1-LR flippee >>>>> Received
Bbw_15 TP901-BP flippee >>>>> Received
Bbw_16 TP901-LR flippee >>>>> Received
Bbw_17 PhiRV1-BP flippee >>>>> Received
Bbw_18 PhiRV1-LR flippee >>>>> Received
oligos design: Freiburg,
parts to get: Bac plasmid:2007 plate3 8EDONE