Difference between revisions of "Part:BBa K2271067:Design"

 
 
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===Design Notes===
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===Part design===
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Mutagenesis
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<p align="justify">
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  This part was designed in the course of a random mutagenesis approach. With the predicted three dimensional strucutre of the TPR domains we carried out our workflow in order to obtain the PEX5 variant we wanted.
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+
  <br>
 +
  After gene synthesis we performed golden gate cloning and made it compatible to the biobrick standard via overhang PCR with the appropriate primers. This product then was ligated into the biobrick backone <b>pSB1C3</b>.
 +
</p>
  
 
===Source===
 
===Source===
  
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ATGGACGTAGGCTCTTGCAGCGTTGGGAACAACCCGCTGGCTCAGTTGCACAAGCACACTCAACAGAATAAGTCTTTGCAGTTCAATCAAAAAAACAACGGCCGTCTAAATGAATCCCCGCTACAGGGGACGAATAAGCCCGGCATCTCAGAGGCGTTTATCTCCAACGTAAATGCCATCTCACAAGAGAACATGGCTAATATGCAAAGGTTCATAAATGGGGAACCCTTGATTGATGATAAGCGTCGTATGGAGATAGGACCGTCTTCTGGGAGACTTCCTCCTTTTAGTAATGTACATTCCTTACAAACCTCCGCTAACCCTACTCAGATAAAGGGCGTCAATGATATAAGCCACTGGAGTCAGGAATTTCAGGGAAGCAACTCAATTCAGAACAGGAATGCCGATACGGGTAATTCCGAGAAAGCCTGGCAGAGAGGCTCAACGACTGCGAGTTCACGTTTTCAGTATCCGAATACTATGATGAACAATTACGCCTATGCAAGTATGAACTCACTTTCCGGTTCCCGTTTGCAGTCACCGGCCTTCATGAATCAACAGCAGAGCGGGAGAAGTAAAGAAGGTGTGAATGAGCAAGAACAGCAACCTTGGACCGACCAATTCGAGAAGTTGGAAAAAGAGGTCAGTGAAAACCTGGACATCAACGACGAAATCGAGAAAGAAGAAAATGTCTCTGAAGTAGAACAGAATAAGCCCGAGACTGTTGAAAAGGAGGAGGGAGTCTACGGCGACCAATACCAGTCTGATTTCCAAGAGGTATGGGACTCAATTCACAAGGACGCGGAAGAGGTCTTACCATCAGAACTGGTAAACGATGACCTAAACCTAGGCGAGGACTATCTAAAATATTTAGGCGGGCGTGTGAATGGCAACATTGAGTATGCCTTTCAATCTAATAATGAGTATTTTAACAACCCAAATGCATATAAAATAGGCTGCCTATTAATGGAAAATGGCGCAAAACTAAGCGAAGCCGCACTTGCGTTCGAAGCGGCCGTCAAGGAGAAGCCAGATCATGTAGACGCCTGGTTAAGATTAGGATTGGTGCAAACGCAAAATCACAAAACTCTAAATGGTATATCCGCACTAGAGGAATGTTTAAAACTGGATCCTAAAAACTTGGAGGCCATGAAGACTTTAGCTATATCTTACATCAACGAAGGATATGACATGAGCGCATTTACAATGTTAGATAAGTGGGCTGAAACAAAATACCCGGAAATATGGTCCAGGATCAAACAGCAAGACGATAAGTTTCAAAAGGAGAAGGGTTTCACACACATCGACATGAATGCCCACATCACAAAGCAATTCTTGCAGCTAGCCAACAACTTATCCACGATAGACCCTGAAATACAACTATGTTTAGGATTGCTATTTGCTACCAAGGATGATTTCGATAAAACAATAGACTGCTTCGAAAGCGCTCTGCGTGTGAATCCCAATGACGAACTTATGTGGAATCGTCTGGGCGCTTCACTTGCGAACTCTAACCGTTCAGAAGAGGCTATACAGGCCTACCACAGGGCTCTACAGTTAAAGCCCAGCTTCGTCCGTGCAAGATACAATCTAGCGGTATCATCTATGAACATTGGATGCTTCAAAGAGGCGGCCGGCTACTTGCTTTCCGTACTAAGCATGCATGAAGTTAATACCAACAACAAAAAAGGTGACGTAGGTTCACTTCTTAACACGTATAACGACACGGTGATCGAAACCCTGAAAAGGGTCTTCATCGCGATGAATCGTGATGACCTACTACAAGAAGTAAAACCTGGGATGGACTTAAAGCGTTTCAAAGGAGAATTTTCATTTTAA
+
<p align="justify">
 +
  The original sequence was taken from NCBI (GenBank: KZV12481.1). After mutagenesis we performed the IDT codon optimization for <i>Saccharomyces cerevisiae</i>, removed any restrictions sites that would interfere with the biobrick or golden gate standard and synthesized the gene with appropiate Golden Gate overhangs.
 +
</p>
  
 
===References===
 
===References===

Latest revision as of 10:25, 29 October 2017


PEX5 variant R19


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 1341
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 1128
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 1633
  • 1000
    COMPATIBLE WITH RFC[1000]


Part design

This part was designed in the course of a random mutagenesis approach. With the predicted three dimensional strucutre of the TPR domains we carried out our workflow in order to obtain the PEX5 variant we wanted.
After gene synthesis we performed golden gate cloning and made it compatible to the biobrick standard via overhang PCR with the appropriate primers. This product then was ligated into the biobrick backone pSB1C3.

Source

The original sequence was taken from NCBI (GenBank: KZV12481.1). After mutagenesis we performed the IDT codon optimization for Saccharomyces cerevisiae, removed any restrictions sites that would interfere with the biobrick or golden gate standard and synthesized the gene with appropiate Golden Gate overhangs.

References