Part:BBa_K3373012:Design
BSR_atzA-HA_atzB-HA_atzC-HA
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 1346
Illegal EcoRI site found at 4254
Illegal EcoRI site found at 7183
Illegal PstI site found at 2990
Illegal PstI site found at 3554 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 1346
Illegal EcoRI site found at 4254
Illegal EcoRI site found at 7183
Illegal NheI site found at 450
Illegal PstI site found at 2990
Illegal PstI site found at 3554 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 1346
Illegal EcoRI site found at 4254
Illegal EcoRI site found at 7183
Illegal XhoI site found at 8887
Illegal XhoI site found at 8995 - 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 1346
Illegal EcoRI site found at 4254
Illegal EcoRI site found at 7183
Illegal PstI site found at 2990
Illegal PstI site found at 3554 - 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 1346
Illegal EcoRI site found at 4254
Illegal EcoRI site found at 7183
Illegal PstI site found at 2990
Illegal PstI site found at 3554
Illegal NgoMIV site found at 2418
Illegal NgoMIV site found at 2818
Illegal NgoMIV site found at 3659
Illegal NgoMIV site found at 5136
Illegal NgoMIV site found at 6173
Illegal NgoMIV site found at 6588
Illegal NgoMIV site found at 9283 - 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI.rc site found at 3725
Illegal SapI.rc site found at 6654
Illegal SapI.rc site found at 9349
Design Notes
This part is a linear DNA that is the result of the digestion of the level M plasmid by the type IIS BsaI restriction enzyme. It is this linear DNA that is integrated into the genomic DNA of Chlamydomonas reinhardtii during the transformation.
Source
The atzA (K3373001); atzB (K3373002) ; atzC (K3373003) enzymes come from the predicted sequence from the Pseudomonas sp. ADP strain genome sequencing.
All the other parts comes from the Chlamydomonas reinhardtii MoClo Kit (Crozet et al. 2018, de Carpentier et al. 2020).
References
1. Crozet P, Navarro FJ, Willmund F, Mehrshahi P, Bakowski K, Lauersen KJ, Pérez-Pérez ME, Auroy P, Gorchs Rovira A, Sauret-Gueto S, Niemeyer J, Spaniol B, Theis J, Trösch R, Westrich LD, Vavitsas K, Baier T, Hübner W, de Carpentier F, Cassarini M, Danon A, Henri J, Marchand CH, de Mia M, Sarkissian K, Baulcombe DC, Peltier G, Crespo JL, Kruse O, Jensen PE, Schroda M, Smith AG, Lemaire SD. Birth of a Photosynthetic Chassis: A MoClo Toolkit Enabling Synthetic Biology in the Microalga Chlamydomonas reinhardtii. ACS Synth Biol. 2018 Sep 21;7(9):2074-2086. doi: 10.1021/acssynbio.8b00251. Epub 2018 Sep 5. PMID: 30165733.
2. Weber E, Engler C, Gruetzner R, Werner S, Marillonnet S. A modular cloning system for standardized assembly of multigene constructs. PLoS One. 2011 Feb 18;6(2):e16765. doi: 10.1371/journal.pone.0016765. PMID: 21364738; PMCID: PMC3041749.
3. de Carpentier F, Le Peillet J, Boisset ND, Crozet P, Lemaire SD, Danon A. Blasticidin S Deaminase: A New Efficient Selectable Marker for Chlamydomonas reinhardtii. Front Plant Sci. 2020 Mar 5;11:242. doi: 10.3389/fpls.2020.00242. PMID: 32211000; PMCID: PMC7066984.
4. de Souza ML, Wackett LP, Boundy-Mills KL, Mandelbaum RT, Sadowsky MJ. Cloning, characterization, and expression of a gene region from Pseudomonas sp. strain ADP involved in the dechlorination of atrazine. Appl Environ Microbiol. 1995 Sep;61(9):3373-8. doi: 10.1128/AEM.61.9.3373-3378.1995. PMID: 7574646; PMCID: PMC167616.
5. Seffernick JL, Aleem A, Osborne JP, Johnson G, Sadowsky MJ, Wackett LP. Hydroxyatrazine N-ethylaminohydrolase (AtzB): an amidohydrolase superfamily enzyme catalyzing deamination and dechlorination. J Bacteriol. 2007 Oct;189(19):6989-97. doi: 10.1128/JB.00630-07. Epub 2007 Jul 27. PMID: 17660279; PMCID: PMC2045197.
6. Sadowsky MJ, Tong Z, de Souza M, Wackett LP. AtzC is a new member of the amidohydrolase protein superfamily and is homologous to other atrazine-metabolizing enzymes. J Bacteriol. 1998 Jan;180(1):152-8. doi: 10.1128/JB.180.1.152-158.1998. PMID: 9422605; PMCID: PMC106861.