Difference between revisions of "Part:BBa K4390002:Design"
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BsmBI recognition site :: BsaI recognition site :: JUMP 5’ fusion site :: Part sequence :: JUMP 3’ fusion site :: BsaI recognition site :: BsmBI recognition site | BsmBI recognition site :: BsaI recognition site :: JUMP 5’ fusion site :: Part sequence :: JUMP 3’ fusion site :: BsaI recognition site :: BsmBI recognition site | ||
| + | |||
| + | OR | ||
CGTCTCGGTCTCC [JUMP 5’ fusion site] :: Part sequence :: [JUMP 3’ fusion site] :: CGAGACCTGAGACG | CGTCTCGGTCTCC [JUMP 5’ fusion site] :: Part sequence :: [JUMP 3’ fusion site] :: CGAGACCTGAGACG | ||
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Composite parts that were ordered in synthetically were designed and ordered with the correct fusion sites, as if they had been assembled from basic parts. | Composite parts that were ordered in synthetically were designed and ordered with the correct fusion sites, as if they had been assembled from basic parts. | ||
| − | For basic parts in the coding sequence (N, O and C), some extra design considerations took place. Parts that did not end with a C part (everything except C, OC and NOC parts) had all stop codons removed, and nucleotides were either inserted or deleted so that the fusion site would not produce a scar. The overall rule for alignment is that the next codon starts immediately after the fusion site. This means that parts begin with the ATG in the R-N fusion site (AATG), the GCC in the N-O fusion site (AGCC) produces an alanine as a scar and the TCG in the O-C (TTCG) fusion site produces serine as a scar. What we often did was remove some nucleotides or codons, and then the scar would regenerate nucleotides or codons that were there before, so less amino acids would be inserted in the composite product. | + | For basic parts in the coding sequence (N, O and C), some extra design considerations took place. Parts that did not end with a C part (everything except C, OC and NOC parts) had all stop codons removed, and nucleotides were either inserted or deleted so that the fusion site would not produce a scar. The overall rule for alignment is that the next codon starts immediately after the fusion site. This means that parts begin with the ATG in the R-N fusion site (AATG), the GCC in the N-O fusion site (AGCC) produces an alanine as a scar and the TCG in the O-C (TTCG) fusion site produces serine as a scar. What we often did was remove some nucleotides or codons, and then the scar would regenerate nucleotides or codons that were there before, so less amino acids would be inserted in the composite product. |
| − | + | ||
| − | + | ||
| − | + | ||
===Source=== | ===Source=== | ||
Revision as of 18:00, 4 September 2022
ArsR expression construct
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 11
Illegal NheI site found at 34 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 196
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
This part was designed with JUMP assembly (a Type IIS assembly method) in mind. All basic parts were designed to be ordered with flanking BsaI and BsmBI sites, as well as the JUMP fusion sites. When basic parts were being ordered in, they would follow the general structure of
BsmBI recognition site :: BsaI recognition site :: JUMP 5’ fusion site :: Part sequence :: JUMP 3’ fusion site :: BsaI recognition site :: BsmBI recognition site
OR
CGTCTCGGTCTCC [JUMP 5’ fusion site] :: Part sequence :: [JUMP 3’ fusion site] :: CGAGACCTGAGACG
| Part type | 5’ Fusion site | 3’ Fusion site |
|---|---|---|
| P (Promoter) | GGAG | TACT |
| R (Ribosome Binding Site) | TACT | AATG |
| N (N-terminus) | AATG | AGCC |
| O (Open Reading Frame) | AGCC | TTCG |
| C (C-terminus) | TTCG | GCTT |
| T (Terminator) | GGCT | CGCT |
Composite parts that were ordered in synthetically were designed and ordered with the correct fusion sites, as if they had been assembled from basic parts.
For basic parts in the coding sequence (N, O and C), some extra design considerations took place. Parts that did not end with a C part (everything except C, OC and NOC parts) had all stop codons removed, and nucleotides were either inserted or deleted so that the fusion site would not produce a scar. The overall rule for alignment is that the next codon starts immediately after the fusion site. This means that parts begin with the ATG in the R-N fusion site (AATG), the GCC in the N-O fusion site (AGCC) produces an alanine as a scar and the TCG in the O-C (TTCG) fusion site produces serine as a scar. What we often did was remove some nucleotides or codons, and then the scar would regenerate nucleotides or codons that were there before, so less amino acids would be inserted in the composite product.
Source
J23100 Promoter - https://parts.igem.org/Part:BBa_J23100 B0034 RBS - https://parts.igem.org/Part:BBa_B0034 arsR Transcriptional Repressor - https://parts.igem.org/Part:BBa_J15101 L2U2H09 Terminator - https://parts.igem.org/Part:BBa_K4390001