BBa_K1975004 1 PhaA-CFP Beta-Ketothiolase fused to Cyan Fluorescent Protein 2016-10-04T11:00:00Z 2016-10-05T06:43:43Z The genetic sequence for PhaA is original from Ralstonia eutropha genomic DNA, but has been optmiized to the expression of B. subtilis. The sequence for CFP genomic DNA from Acropora aculeus. This sequence has also been optimized for expression in B. subtilis. This is a propionate-CoA transferase (commonly known as PhaA). This enzyme catalyzes the first step in the production of PHB. The reaction it catalyzes is two molecules of acetyl-CoA is being putting together to one molecule of acetoacetyl-CoA under the usage of a propanoate that will be converted to acetate. By combining it with the synthetic gene, LPE it can be used in the synthesis of P(LA-co-3HB). This gene is fused with the Cyan Fluorescence Protein with a small linker of 6 bp between. The fused CFP is used to visual the expression of the protein. Hereby, the job will be easier to screen for positive transformants after induction (if such is used as in our case). The sequence for PhaA is taken from the BioBrick Part:BBa_K759004 and is optimized to be expressed in Bacillus subtilis. false false _2442_ 30862 30862 9 false To secure the optimal expression we choose to make a codon optimization of the genes. This was done through the Codon Optimization tool at IDT's webpage. Besides that we looked through the literature to figure our if the PhaA and CFP should be directly attached or if they should have some kind of linker. After looking through different studies that have been working with PhaA in Escherichia coli we came to the conclusion that a linker should not be necessary to secure high expression and activity. We therefore decided to only make a small linker (6 bp) between the gene and the tag. false Joachim Steen Larsen component2486164 1 BBa_K1975005 component2486163 1 BBa_K1975002 annotation2486164 1 BBa_K1975005 range2486164 1 1189 1893 annotation2486163 1 BBa_K1975002 range2486163 1 1 1182 BBa_K1975005 1 BBa_K1975005 Cyan Fluorescent Protein 2016-10-04T11:00:00Z 2016-10-05T06:40:52Z The sequence for CFP genomic DNA from Acropora aculeus. This sequence has also been optimized for expression in B. subtilis. This is Cyan Fluorescent Protein that can be used to tag proteins of interest. It is a fluorescent protein that will show a blue color when it is excited at 405nm and it has the highest absorbance at 485nm. This sequence has been used to tag PhaA to visualize the expression of the protein. If the protein was expressed correctly, the cells would show a blue color when excited. The sequence is optimized for use in Bacillus subtilis. false false _2442_ 30862 30862 9 false To secure the optimal expression we choose to make a codon optimization of the genes. This was done through the Codon Optimization tool at IDT's webpage. false Joachim Steen Larsen BBa_K1975002 1 BBa_K1975002 PhaA - Beta-kethothiolase 2016-06-27T11:00:00Z 2016-06-28T04:11:00Z Genomic DNA Will be coming later. false false _2442_ 30862 30862 9 false Codon optimized for B. subtilis. false Joachim Steen Larsen BBa_K1975002_sequence 1 atgacggacgtggtcattgtatccgctgccagaactgccgtcggaaagttcggtggaagccttgcaaaaattccagcgccggaacttggggcggtcgtgataaaagcggcactggaacgcgcaggggtaaagccagagcaagtgtccgaggtaataatgggccaggtccttaccgcagggtcaggtcaaaaccctgctcgccaagccgcaatcaaggcggggctgcctgcgatggttccagcaatgacgatcaacaaggtatgcggctcaggcttgaaagcagttatgttggccgcaaatgcgattatggctggagatgcggagatagttgttgctggaggccaggagaatatgtcagccgccccgcatgtactcccaggaagccgggacgggtttcgtatgggtgatgctaagctggtcgatacaatgatagtagatggactgtgggacgtctacaaccaatatcatatgggcataaccgccgagaacgtcgcaaaggaatatggaatcactcgcgaggctcaagacgagtttgccgtgggatctcagaataaagccgaggcggcccaaaaagctggcaagtttgatgaagaaatagttccggttttaatacctcaacgcaaaggtgatcctgtggcatttaaaacggatgagtttgtccgccagggagcaacgcttgatagtatgtcagggttgaagccagcgtttgacaaagctggcaccgtcactgctgccaacgctagtggactcaatgatggggcggctgcggtggtcgtcatgagtgctgccaaggcgaaggagcttgggctcaccccgctggccactataaaaagttatgctaacgcgggagtcgaccctaaggttatgggcatgggaccagttccggcctctaaaagagcattgagcagagctgaatggacaccgcaggaccttgaccttatggagatcaatgaagccttcgcagctcaagccctggcagtccatcagcagatggggtgggatacttctaaagtaaacgtgaatggtggtgcgatcgcaattgggcatccaataggcgcgagcggctgtagaattttagtcactctgttacacgaaatgaagagacgtgatgcaaaaaagggcttagcgtcattgtgcatagggggaggtatgggggtcgcgttagcagtagaaagaaaatac BBa_K1975005_sequence 1 atgtctctttctaaacacgggataacccaggaaatgccgacaaagtatcacatgaaagggtctgtgaatggacacgaatttgagatcgaaggtgttggcaccggccacccatacgaaggcacccatatggccgaattggttattatcaagcccgctggcaagccattaccgttttcctttgacattctgtctacagtaattcaatatggaaatagatgcttcacaaagtatcctgccgatctgccggattactttaaacaggcgtaccctggaggtatgagctatgaacgctccttcgtctaccaggatggcggtatagcgacggctagttggaacgtcggtctggagggcaattgcttcatacacaagtctacttatctgggcgtcaacttcccagctgatggtcctgtaatgacgaaaaagaccatagggtgggacaaagcgtttgagaaaatgacagggtttaacgaggtgcttagaggggatgtaaccgaatttctcatgctggagggtggggggtatcatagctgccagtttcattcaacctacaaacctgagaaaccagtagagttacctccaaatcatgtgatagaacaccacatagtgcgcaccgatttaggcaagaccgcaaagggctttatggttaaacttgtccaacacgcagcagcacatgttaatccgttgaaagtgcaagcggccgcgtaa BBa_K1975004_sequence 1 atgacggacgtggtcattgtatccgctgccagaactgccgtcggaaagttcggtggaagccttgcaaaaattccagcgccggaacttggggcggtcgtgataaaagcggcactggaacgcgcaggggtaaagccagagcaagtgtccgaggtaataatgggccaggtccttaccgcagggtcaggtcaaaaccctgctcgccaagccgcaatcaaggcggggctgcctgcgatggttccagcaatgacgatcaacaaggtatgcggctcaggcttgaaagcagttatgttggccgcaaatgcgattatggctggagatgcggagatagttgttgctggaggccaggagaatatgtcagccgccccgcatgtactcccaggaagccgggacgggtttcgtatgggtgatgctaagctggtcgatacaatgatagtagatggactgtgggacgtctacaaccaatatcatatgggcataaccgccgagaacgtcgcaaaggaatatggaatcactcgcgaggctcaagacgagtttgccgtgggatctcagaataaagccgaggcggcccaaaaagctggcaagtttgatgaagaaatagttccggttttaatacctcaacgcaaaggtgatcctgtggcatttaaaacggatgagtttgtccgccagggagcaacgcttgatagtatgtcagggttgaagccagcgtttgacaaagctggcaccgtcactgctgccaacgctagtggactcaatgatggggcggctgcggtggtcgtcatgagtgctgccaaggcgaaggagcttgggctcaccccgctggccactataaaaagttatgctaacgcgggagtcgaccctaaggttatgggcatgggaccagttccggcctctaaaagagcattgagcagagctgaatggacaccgcaggaccttgaccttatggagatcaatgaagccttcgcagctcaagccctggcagtccatcagcagatggggtgggatacttctaaagtaaacgtgaatggtggtgcgatcgcaattgggcatccaataggcgcgagcggctgtagaattttagtcactctgttacacgaaatgaagagacgtgatgcaaaaaagggcttagcgtcattgtgcatagggggaggtatgggggtcgcgttagcagtagaaagaaaatactactagatgtctctttctaaacacgggataacccaggaaatgccgacaaagtatcacatgaaagggtctgtgaatggacacgaatttgagatcgaaggtgttggcaccggccacccatacgaaggcacccatatggccgaattggttattatcaagcccgctggcaagccattaccgttttcctttgacattctgtctacagtaattcaatatggaaatagatgcttcacaaagtatcctgccgatctgccggattactttaaacaggcgtaccctggaggtatgagctatgaacgctccttcgtctaccaggatggcggtatagcgacggctagttggaacgtcggtctggagggcaattgcttcatacacaagtctacttatctgggcgtcaacttcccagctgatggtcctgtaatgacgaaaaagaccatagggtgggacaaagcgtttgagaaaatgacagggtttaacgaggtgcttagaggggatgtaaccgaatttctcatgctggagggtggggggtatcatagctgccagtttcattcaacctacaaacctgagaaaccagtagagttacctccaaatcatgtgatagaacaccacatagtgcgcaccgatttaggcaagaccgcaaagggctttatggttaaacttgtccaacacgcagcagcacatgttaatccgttgaaagtgcaagcggccgcgtaa igem2sbol 1 iGEM to SBOL conversion Conversion of the iGEM parts registry to SBOL2.1 Chris J. Myers James Alastair McLaughlin 2017-03-06T15:00:00.000Z