Difference between revisions of "Part:BBa K2570001:Design"
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Mutation of restriction sites EcoRⅠ, SpeⅠand PstⅠ. | Mutation of restriction sites EcoRⅠ, SpeⅠand PstⅠ. | ||
| − | + | <span style="font-weight:bold;">Site-directed mutation site </span> | |
Mutation 1: Aro10-Pst1 （Pro 619-621）cctgca-ccggca | Mutation 1: Aro10-Pst1 （Pro 619-621）cctgca-ccggca | ||
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===Source=== | ===Source=== | ||
| − | The phenylpyruvate decarboxylase from S. cerevisiae (Aro10, NP 010668.3). | + | The phenylpyruvate decarboxylase from ''S. cerevisiae'' (Aro10, NP 010668.3). |
https://www.ncbi.nlm.nih.gov/protein/NP_010668.3?report=genbank&log$=protalign&blast_rank=1&RID=V0V04K3B015 | https://www.ncbi.nlm.nih.gov/protein/NP_010668.3?report=genbank&log$=protalign&blast_rank=1&RID=V0V04K3B015 | ||
Latest revision as of 04:37, 10 October 2018
Aro10
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
We ordered the part DNA from a synthesis company and had codon optimization of the sequence. Mutation of restriction sites EcoRⅠ, SpeⅠand PstⅠ.
Site-directed mutation site
Mutation 1: Aro10-Pst1 （Pro 619-621）cctgca-ccggca
Sense primer: TATATTTTTGTTCCGGCAGATTTTG
Anti-sense primer: AATCTGCCGGAACAAAAATATAACC
Mutation 2: Aro10-Spe1 （Thr 741-744）actagt- accagt
Sense primer: ACAAGATTACCAGTTGGATATATTC
Anti-sense primer: ATATATCCAACTGGTAATCTTGTTG
Mutation 3: Aro10-EcoR1（Asn 1791-1794）gaattc-gaactc
Sense primer: CTTAAGAACTCAAACAAAAGAAGCG
Anti-sense primer: TTTGTTTGAGTTCTTAAGCTCCTCC
Source
The phenylpyruvate decarboxylase from S. cerevisiae (Aro10, NP 010668.3).
References
Wang, P., Yang, X., Lin, B., Huang, J., & Tao, Y. (2017). Cofactor self-sufficient whole-cell biocatalysts for the production of 2-phenylethanol. Metabolic Engineering, 44(August), 143–149.