BBa_K2013017 1 BBa_K2013017 2-oxopent-4-enoate hydratase 2016-10-11T11:00:00Z 2016-10-15T08:16:58Z This sequence is from Pseudomonas putida F1 This part contains the sequence of coding 2-oxopent-4-enoate hydratase which catalyzes 2-hydroxymuconate semialdehyde into 2-oxopent-4-enoate.In addition，We carried out codon optimization on this part Experimental Validation This part is validated through Four ways: amplification, PCR, Enzyme cutting and Sequence. Amplification Enzyme:KOD Primer-F:5GAATTCGCGGCCGCTTCTAGAGTACTAGAGAAAGAGGAGAAATACTAGATGACCAACGTGAACG-3′ Primer-R:5′- CGCTACTAGTATTATTACCACGCGGTGTGATACAG-3′ Results PCR Enzyme:Taq Primer-F:5′-CCACCTGACGTCTAAGAAAC-3′ Primer-R:5′-GTATTACCGCCTTTGAGTGA-3′ Results Double digestion After the assembly ,the plasmid was transferred into the Competent E. coli top10. After culturing overnight in LB,we minipreped the plasmid for double digestion .The first cutting procedure was performed with EcoRI and NcoI restriction endonuclease. The second cutting procedure was performed with PstI and NcoI restriction endonuclease.The plasmid was cutted in a 25μL system at 37 ℃ for 1 hours. The Electrophoresis was performed on a 1％ Agar Results false false _2480_ 32794 32794 9 false This is a Translational units part that encodes 2-oxopent-4-enoate hydratase false Mei Deng annotation2496410 1 2-hydroxy-6-oxohepta-2,4-dienoate hydrolase range2496410 1 20 852 annotation2496052 1 RBS range2496052 1 1 13 BBa_K2013017_sequence 1 aaagaggagaaatactagatgaccaacgtgaacgcggagatcggccgtatggttctggcgggtggcattgaaaccaacctgcacgacgtgggtgcgggtaacccggtggttctggtgcacggtagcggtccgggtgttaccgcgtgggcgaactggcgtaccgttatgccggagctgagccgtcaccgtcgtgtgatcgcgccggacatggttggcttcggttttacccagcgtccgcacggcatccactacggtgtggagagctgggttgcgcacctggcgggcattctggatgcgctggaactggaccgtgtggatctggttggtaacagctttggtggcgcgctgagcctggcgttcgcgattcgttttccgcaccgtgtgcgtcgtctggttctgatgggcgcggtgggtgttagctttgagctgaccgacggcctggatgcggtgtggggttacgaaccgagcgtgccgaacatgcgtaaggttatggactacttcgcgtatgatcgtagcctggttagcgatgagctggcggaactgcgttataaagcgagcacccgtccgggtttccaagaggcgtttgcgagcatgttcccggcgccgcgtcagcgttgggtggacgcgctggcgagcagcgaccaagatatccgtgatattcgtcacgaaaccctgatcctgcacggtcgtgacgatcgtgttattccgctggaaaccagcctgcgtctgaaccagctgatcgaaccgagccaactgcacgtgttcggccgttgcggtcactgggttcagatcgagcaaaaccgtggttttattcgtctggtgaacgacttcctggcggcggaagattaataa 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