LOCUS       sequence-349663-        6821 bp    DNA     circular SYN 14-APR-2025
DEFINITION  expresses a fatty alcohol oxidase in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6821)
  AUTHORS   Rembeza E, Boverio A, Fraaije MW, Engqvist MKM
  TITLE     Discovery of Two Novel Oxidases Using a High-Throughput Activity 
            Screen.
  JOURNAL   Chembiochem. 2022 Jan 19;23(2):e202100510. doi: 
            10.1002/cbic.202100510. Epub 2021 Nov 18.
  PUBMED    34709726
REFERENCE   2  (bases 1 to 6821)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Chembiochem."; date: "2022-01-19"; volume: "23(2):e202100510. doi";
            pages: " 10.1002/cbic.202100510. Epub 2021 Nov 18"
COMMENT     Sequence Label: A0A075HNX4
FEATURES             Location/Qualifiers
     source          1..6821
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             68..97
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     primer_bind     complement(150..168)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      164..211
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      248..703
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(335..354)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     545..566
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        729..833
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(834..902,903..1694)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1052..1071)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1865..2453
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2354..2373
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2607..2624
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2639..2778
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2704..2723)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2864..2886
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2880..3071)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3880..4962)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     4924..4943
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4963..5040)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5246..5265
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5349..5368
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5349..5367
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5368..5392
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             5407..5429
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 tgattggaca actgcactgc ttgaggctga tggtcaggtg ggcgattttc ctgcagcagc
       61 tctagaacac caccatcacc accaccacca ccaccactga gatccggctg ctaacaaagc
      121 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
      181 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc
      241 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
      301 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
      361 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
      421 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
      481 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
      541 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
      601 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
      661 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
      721 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
      781 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
      841 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
      901 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
      961 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     1021 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     1081 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     1141 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     1201 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     1261 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     1321 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag
     1381 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     1441 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     1501 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     1561 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     1621 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     1681 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     1741 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     1801 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     1861 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     1921 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     1981 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     2041 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     2101 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     2161 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     2221 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     2281 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     2341 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     2401 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     2461 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     2521 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     2581 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     2641 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct
     2701 cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt
     2761 gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     2821 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     2881 cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg
     2941 tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc
     3001 tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc
     3061 tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg
     3121 atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg
     3181 gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact
     3241 cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag
     3301 catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga
     3361 ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg
     3421 cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg
     3481 caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg
     3541 gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg
     3601 acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc
     3661 gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt
     3721 cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc
     3781 gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt
     3841 gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg
     3901 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     3961 cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc
     4021 cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag
     4081 gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc
     4141 atcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg
     4201 cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc
     4261 attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc
     4321 cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg
     4381 cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac
     4441 cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg
     4501 tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc
     4561 aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag
     4621 aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac
     4681 cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc
     4741 gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg
     4801 ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg
     4861 cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac
     4921 accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg
     4981 actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc
     5041 cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt
     5101 tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca
     5161 gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga
     5221 agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac
     5281 ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc
     5341 cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt cccctctaga
     5401 aataattttg tttaacttta agaaggagat atacatatgg cacaaggcgc ccagaggaag
     5461 aatttcggcc acaatcagat ccttagacct tctgcagctt atacaccggt tgatgaacaa
     5521 gaggtactcc agatcttgga tcgacacaga ggacagagga tacgagctgt gggacgattg
     5581 cattcctggt ccgaggcagt taccggtgat ggtgtacttt tggacttaca gagactaaat
     5641 gatgtcagac tgcagtcgga cggtgaccaa ctggtcgcaa cggtgggcgc tggttgtcag
     5701 attaaaagac ttttgaagga gttgaatcgt gaaggagcaa ccttacactc tctgggtctc
     5761 atcacagctc agactattgc cggagctatt tccacaggta cacatggatc cggacgtaac
     5821 tccatgtctc attacgttgt tggtgtgcga ttggcatgtt acgatgcttc gacaggtcag
     5881 gctattatcg aggagttgtc tgcaggagaa cctcttcagg ctgccagatg cagtttaggt
     5941 tccttgggca tcatactggc agtaagaatt cgttgtcgag aacaatacaa cgtacaggag
     6001 catttcacgg agtcacgaag gctgttagac gttatggacg ctgaggctcc ttttccactt
     6061 caacagtttt acttgttgcc gtggagatgg agttacttta tccagcacag aagagaggac
     6121 gataggccaa gatcgaggct agctagacta tatagattgt actggttggg aactatggat
     6181 tatggtttga tattacagat cttgtttctt gagagagtcg cccgatcaag aagattaatc
     6241 agactggctt tcagacgtat cattcctgct ttcttgatta gaaattggag agttactgac
     6301 cgttcgtctt ccatgcttgt catgagacat gacgctttca ggcacatcga aattgagttg
     6361 tttgtaagaa gagaccaact cgcagacgca ttgggattta cccaagaagt aattaagatt
     6421 gctggtggtc gtgaatccgc attgagtgct gacaatcaga gaagaatcga ggagctcggt
     6481 atgcaagagg ccctagctgg tttgcatgac cagtattgtc atcacttccc aatctgcgtt
     6541 cgtagagttt tgccggacga caccctaata agcatggctt ccggtgctgg cgaagattgg
     6601 tacgctttgt ctttcatttc atacgccaaa ccggcaagac gtgccgggtt ctctcttttc
     6661 gccagtttca tggctagatc aatgtcgaga cttttccacg ctaggcccca ttggggaaag
     6721 gtttgtccat tggaagccga tgagctaact agtttgtatc cacgtttcga tgcctttagg
     6781 actgtttgta atacactaga cccacagggt gttttccaaa a
//
