Difference between revisions of "Part:BBa K863101:Design"
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__NOTOC__ | __NOTOC__ | ||
<partinfo>BBa_K863101 short</partinfo> | <partinfo>BBa_K863101 short</partinfo> | ||
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===Design Notes=== | ===Design Notes=== | ||
− | The sequence starts with the Freiburg-prefix (ATG | + | The sequence starts with the rest of the Freiburg-prefix (ATG and ''Ngo''MIV-restriction-site). The A from the startcodon ATG is the last base of the ''Xba''I restriction-site of the [https://parts.igem.org/Assembly_standard_25 Standard 25 Prefix (Read here)]. I isolated 12 conserved Bases (4 AS) upstream of the cellulose binding domain and 9 bases downstream of the exoglucanase gene and added a short N-terminal (Glycine, Serine) linker. The sequence ends with the ''Age''I-restriction-site of the Freiburg-Suffix, to make in frame assembly possible. |
− | + | ||
+ | Primers that were used to make this BioBrick: | ||
+ | {| | ||
+ | |CBDcex_Freiburg-Prefix||GCTAGAATTCGCGGCCGCTTCTAGATGGCCGGCGGTCCGGCCGGGTGCCAGGTG | ||
+ | |- | ||
+ | |CBDcex_2AS-Link_Frei||CTGCAGCGGCCGCTACTAGTATTAACCGGTGCTGCCGCCGACCGTGCAGGGCGTGC | ||
+ | |} | ||
===Source=== | ===Source=== | ||
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===References=== | ===References=== | ||
+ | '''Contribution (Waterloo iGEM 2020)''' | ||
+ | |||
+ | McLean, B.W., Bray, M.R., Boraston, A.B., Gilkes, N.R., Haynes, C.A., & Kilburn, D.G. (2000). Analysis of binding of the family 2a carbohydrate -binding module from ''Cellulomonas fimi'' xylanase 10A to cellulose: specificity and identification of functionally important amino acid residues. ''Protein Engineering'', ''13''(11). 801-809. | ||
+ | |||
+ | Rodriguez, B., Kavoosi, M., Koska, J., Creagh, A.L., Kilburn, D.G., & Haynes, C.A. (2004). Inexpensive and Generic Affinity Purification of Recombinant Proteins Using a Family 2a CBM Fusion Tag. ''Biotechnology, 20.'' 1479 - 1489. |
Latest revision as of 02:57, 22 October 2020
Cellulose binding Domain of Cellulomonas Fimi Exoglucanse (Freiburg-Standard)
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 4
Illegal AgeI site found at 337 - 1000COMPATIBLE WITH RFC[1000]
Design Notes
The sequence starts with the rest of the Freiburg-prefix (ATG and NgoMIV-restriction-site). The A from the startcodon ATG is the last base of the XbaI restriction-site of the Standard 25 Prefix (Read here). I isolated 12 conserved Bases (4 AS) upstream of the cellulose binding domain and 9 bases downstream of the exoglucanase gene and added a short N-terminal (Glycine, Serine) linker. The sequence ends with the AgeI-restriction-site of the Freiburg-Suffix, to make in frame assembly possible.
Primers that were used to make this BioBrick:
CBDcex_Freiburg-Prefix | GCTAGAATTCGCGGCCGCTTCTAGATGGCCGGCGGTCCGGCCGGGTGCCAGGTG |
CBDcex_2AS-Link_Frei | CTGCAGCGGCCGCTACTAGTATTAACCGGTGCTGCCGCCGACCGTGCAGGGCGTGC |
Source
Cloning-Vector with a CBDcex-Barnase fusion-protein
References
Contribution (Waterloo iGEM 2020)
McLean, B.W., Bray, M.R., Boraston, A.B., Gilkes, N.R., Haynes, C.A., & Kilburn, D.G. (2000). Analysis of binding of the family 2a carbohydrate -binding module from Cellulomonas fimi xylanase 10A to cellulose: specificity and identification of functionally important amino acid residues. Protein Engineering, 13(11). 801-809.
Rodriguez, B., Kavoosi, M., Koska, J., Creagh, A.L., Kilburn, D.G., & Haynes, C.A. (2004). Inexpensive and Generic Affinity Purification of Recombinant Proteins Using a Family 2a CBM Fusion Tag. Biotechnology, 20. 1479 - 1489.