LOCUS       sequence-430563-        7982 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses DaPFOR in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7982)
  AUTHORS   Ng TL, Silver PA
  TITLE     Sustainable B(12)-Dependent Dehalogenation of Organohalides in E. 
            coli.
  JOURNAL   ACS Chem Biol. 2024 Feb 16;19(2):380-391. doi: 
            10.1021/acschembio.3c00585. Epub 2024 Jan 22.
  PUBMED    38254247
REFERENCE   2  (bases 1 to 7982)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/acschembio.3c00585"; journalName: "ACS Chem Biol"; date: 
            "2024-02-16- 16"; volume: "19"; issue: "2"; pages: "380-391"
COMMENT     Sequence Label: pACYCDUET-StrepSUMO-DaPFOR
FEATURES             Location/Qualifiers
     source          1..7982
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /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)."
     CDS             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        214..232
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    233..257
                     /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)."
     CDS             303..326
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             4338..4382
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(4420..4438)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4434..4481
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(4702..5361)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     5276..5295
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(5362..5464)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(5856..5873)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5990..6535)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(6070..6089)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6760..7842)
                     /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     7804..7823
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7843..7920)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(7966..7982,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agccaggatc cgaattcgag
      121 ctcggcgcgc ctgcaggtcg acaagcttgc ggccgcataa tgcttaagtc gaacagaaag
      181 taatcgtatt gtacacggcc gcataatcga aattaatacg actcactata ggggaattgt
      241 gagcggataa caattcccca tcttagtata ttagttaagt ataagaagga gatatacata
      301 tgtggtctca cccacaattt gagaaaggca gtctgcaaga tagtgaggtg aaccaagagg
      361 cgaaacccga ggtaaaaccc gaagtgaagc ctgagaccca tattaacctc aaagtaagtg
      421 acggttcgag cgaaattttc tttaaaatta agaagacaac cccgcttcgt cgtctgatgg
      481 aggctttcgc caaacgtcaa gggaaagaga tggatagcct gcgcttttta tatgatggta
      541 ttcgaattca agcggatcag gctcctgaag atctggatat ggaagacaat gatatcatag
      601 aggctcatag agaacaaatt ggcggcatgg gaaagaaaat gatgacgact gatggcaata
      661 cagcgacagc gcacgtggcg tatgccatga gcgaagtcgc cgccatctac cccatcaccc
      721 cttcctcgac catgggcgag gaggctgacg actgggcggc gcaaggacgc aagaacatct
      781 ttggccagac cctgaccata cgcgaaatgc agtccgaggc cggcgccgcc ggcgcggtgc
      841 acggggccct ggcggccggc gccctgacca cgaccttcac ggcgtcgcag ggtctgctgc
      901 tgatgatccc caacatgtac aagatctccg gcgaacttct gcccggcgtg ttccacgtca
      961 ccgcccgcgc catcgccgcg cacgccctgt ccatcttcgg tgaccaccag gatatctacg
     1021 ccgcgcgcca gacaggcttc gccatgctcg cctccagctc ggtgcaggag gcccacgaca
     1081 tggccctggt ggcccacttg gcggccatcg agtccaacgt gccgttcatg cacttcttcg
     1141 acggattccg cacctcgcac gaaatccaga agatcgaggt cctggactac gcggacatgg
     1201 cctcgctggt gaaccagaag gccctggcgg aattccgcgc caagtccatg aaccccgagc
     1261 acccccacgt gcgcggcacg gcccagaacc ccgacatcta cttccagggt cgcgaggcag
     1321 ccaaccccta ctacctcaag gtgcccggca tcgttgccga gtacatgcag aaggtcgcct
     1381 ccctcacggg ccgcagctac aagctctttg actacgtggg tgctcccgac gccgagcgcg
     1441 tcatcgtgtc catgggctcc tcgtgcgaga ccatcgagga ggtcatcaac cacctcgcgg
     1501 ccaagggcga aaagatcggc ctgatcaagg tccgcctgta caggcccttc gtaagcgagg
     1561 ccttcttcgc tgctctgccc gcttcggcca aggtcatcac ggtcctcgac cgcaccaagg
     1621 aacccggcgc gcccggcgat ccgctctacc tcgacgtgtg ctcggccttc gtggagcgcg
     1681 gcgaagccat gcccaagatc ctggccggcc gctacggcct gggttccaag gaattcagcc
     1741 cggccatggt caagtccgtg tacgacaaca tgtccggcgc taagaagaac cacttcaccg
     1801 tgggcatcga agacgacgtg accggcactt cgctgccggt ggacaacgcc ttcgccgaca
     1861 ccacgcccaa gggcaccatc cagtgccagt tctggggcct cggcgccgac ggcactgtgg
     1921 gcgccaacaa gcaggccatc aagatcatcg gcgacaacac ggacctgttt gcccagggtt
     1981 acttctccta cgactccaag aaatcgggcg gcatcaccat ctcgcacctg cgcttcggcg
     2041 agaagcccat ccagtccacc tacctggtca acagggccga ctatgtcgcc tgtcacaacc
     2101 cggcctacgt gggcatatac gacatcctcg aaggcatcaa ggatggcgga accttcgtgc
     2161 tcaactcgcc ttggagcagc ctcgaggaca tggacaagca cctgccctcc ggcatcaagc
     2221 gcaccatcgc gaacaagaag ctcaagttct acaacatcga cgcggtgaaa atcgccaccg
     2281 atgtgggact gggcggccgc atcaacatga tcatgcagac ggccttcttc aagctggccg
     2341 gagtgctgcc cttcgaaaag gccgtggatc tgctcaagaa gtccatccac aaggcctacg
     2401 gcaaaaaggg cgagaagatc gtcaagatga acaccgacgc cgtggaccag gccgtcacct
     2461 ccctgcagga attcaagtat ccggattcct ggaaggacgc tcccgctgag accaaggccg
     2521 agcccatgac gaacgagttc ttcaagaacg tcgtcaagcc catcctgacc cagcagggcg
     2581 acaagctgcc ggtgagcgcc ttcgaggccg acggccgttt ccccctcggc accagccagt
     2641 tcgagaagcg cggcgtggcc atcaacgtgc cgcagtgggt ccccgagaac tgcatccagt
     2701 gcaaccagtg cgccttcgtc tgtccgcaca gcgccatcct gcccgtgctg gccaaggaag
     2761 aggagttggt cggcgcgccg gcgaacttca cggccctgga agccaagggc aaggagctca
     2821 agggctacaa gttccgcatc cagatcaaca ccctggactg catgggctgc ggcaactgcg
     2881 ccgacatctg tccgcccaag gaaaaggctc tggtcatgca gcccctggat acccagcgcg
     2941 acgcgcaggt gcccaacctg gagtacgcag cgcgcatccc ggtcaaatcc gaggtgctgc
     3001 cgcgcgactc gctcaagggc agccagttcc aggagcctct catggaattc tcgggcgcct
     3061 gctcgggctg cggcgagacg ccctacgtgc gcgtcatcac ccagctcttc ggcgagcgca
     3121 tgttcattgc caacgccacg ggttgctcgt ccatctgggg cgcgtcggct ccttccatgc
     3181 cttacaagac caaccgcctc ggacaaggcc cggcctgggg taactccctg ttcgaagacg
     3241 cggccgaata cggcttcggc atgaacatgt ccatgttcgc ccgccgcacg catttggccg
     3301 atcttgccgc caaggccctg gagagcgatg cctccggcga tgtcaaggaa gccctgcagg
     3361 gctggcttgc cggcaagaac gatcccatca agtccaagga atacggcgac aagctcaaga
     3421 agctgctggc tggtcagaag gatggtctgc tcggacagat cgccgccatg tccgacctgt
     3481 acaccaagaa gagcgtgtgg atcttcggtg gcgacggctg ggcctacgac atcggttacg
     3541 gcggcctgga ccatgtgctc gcctcgggcg aggacgtgaa cgtcttcgtc atggataccg
     3601 aggtctactc caacaccggc ggccagtcct ccaaggcaac gcccacgggc gccgtggcca
     3661 agttcgcggc ggccggcaag cgtaccggca agaaggacct ggcgcgcatg gtcatgacct
     3721 acggctacgt ctacgtggct acggtctcca tgggttacag caagcagcag ttcctcaagg
     3781 tgctcaagga agccgaaagc ttccccggcc cctcgctggt catcgcctat gctacctgca
     3841 tcaaccaggg tctgcgcaag ggcatgggca agagccagga cgtcatgaac accgcggtca
     3901 agtccggtta ctggccgctg ttccgctacg atccgcgctt ggccgcccag ggcaagaacc
     3961 ccttccagct cgactccaag gctcctgacg gttccgtcga ggagttcctg atggcccaga
     4021 accgcttcgc cgtcctcgat cggtccttcc ccgaggacgc caagagactg cgcgcccagg
     4081 tcgctcacga attggacgtg cgtttcaagg agttggagca catggccgcc acgaacatct
     4141 tcgagtcctt cgcgccagcg ggcggcaagg ccgatggttc ggtggatttc ggcgaaggtg
     4201 cggagttctg cacgcgcgac gatactccca tgatggcccg acctgattcc ggtgaggcct
     4261 gcgaccagaa ccgcgctggc acgagcgaac agcagggaga cctcagcaag cggacgaaga
     4321 agtagctcga gtctggtaaa gaaaccgctg ctgcgaaatt tgaacgccag cacatggact
     4381 cgtctactag cgcagcttaa ttaacctagg ctgctgccac cgctgagcaa taactagcat
     4441 aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaacct caggcatttg
     4501 agaagcacac ggtcacactg cttccggtag tcaataaacc ggtaaaccag caatagacat
     4561 aagcggctat ttaacgaccc tgccctgaac cgacgaccgg gtcgaatttg ctttcgaatt
     4621 tctgccattc atccgcttat tatcacttat tcaggcgtag caaccaggcg tttaagggca
     4681 ccaataactg ccttaaaaaa attacgcccc gccctgccac tcatcgcagt actgttgtaa
     4741 ttcattaagc attctgccga catggaagcc atcacaaacg gcatgatgaa cctgaatcgc
     4801 cagcggcatc agcaccttgt cgccttgcgt ataatatttg cccatagtga aaacgggggc
     4861 gaagaagttg tccatattgg ccacgtttaa atcaaaactg gtgaaactca cccagggatt
     4921 ggctgagacg aaaaacatat tctcaataaa ccctttaggg aaataggcca ggttttcacc
     4981 gtaacacgcc acatcttgcg aatatatgtg tagaaactgc cggaaatcgt cgtggtattc
     5041 actccagagc gatgaaaacg tttcagtttg ctcatggaaa acggtgtaac aagggtgaac
     5101 actatcccat atcaccagct caccgtcttt cattgccata cggaactccg gatgagcatt
     5161 catcaggcgg gcaagaatgt gaataaaggc cggataaaac ttgtgcttat ttttctttac
     5221 ggtctttaaa aaggccgtaa tatccagctg aacggtctgg ttataggtac attgagcaac
     5281 tgactgaaat gcctcaaaat gttctttacg atgccattgg gatatatcaa cggtggtata
     5341 tccagtgatt tttttctcca ttttagcttc cttagctcct gaaaatctcg ataactcaaa
     5401 aaatacgccc ggtagtgatc ttatttcatt atggtgaaag ttggaacctc ttacgtgccg
     5461 atcaacgtct cattttcgcc aaaagttggc ccagggcttc ccggtatcaa cagggacacc
     5521 aggatttatt tattctgcga agtgatcttc cgtcacaggt atttattcgg cgcaaagtgc
     5581 gtcgggtgat gctgccaact tactgattta gtgtatgatg gtgtttttga ggtgctccag
     5641 tggcttctgt ttctatcagc tgtccctcct gttcagctac tgacggggtg gtgcgtaacg
     5701 gcaaaagcac cgccggacat cagcgctagc ggagtgtata ctggcttact atgttggcac
     5761 tgatgagggt gtcagtgaag tgcttcatgt ggcaggagaa aaaaggctgc accggtgcgt
     5821 cagcagaata tgtgatacag gatatattcc gcttcctcgc tcactgactc gctacgctcg
     5881 gtcgttcgac tgcggcgagc ggaaatggct tacgaacggg gcggagattt cctggaagat
     5941 gccaggaaga tacttaacag ggaagtgaga gggccgcggc aaagccgttt ttccataggc
     6001 tccgcccccc tgacaagcat cacgaaatct gacgctcaaa tcagtggtgg cgaaacccga
     6061 caggactata aagataccag gcgtttcccc ctggcggctc cctcgtgcgc tctcctgttc
     6121 ctgcctttcg gtttaccggt gtcattccgc tgttatggcc gcgtttgtct cattccacgc
     6181 ctgacactca gttccgggta ggcagttcgc tccaagctgg actgtatgca cgaacccccc
     6241 gttcagtccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggaaaga
     6301 catgcaaaag caccactggc agcagccact ggtaattgat ttagaggagt tagtcttgaa
     6361 gtcatgcgcc ggttaaggct aaactgaaag gacaagtttt ggtgactgcg ctcctccaag
     6421 ccagttacct cggttcaaag agttggtagc tcagagaacc ttcgaaaaac cgccctgcaa
     6481 ggcggttttt tcgttttcag agcaagagat tacgcgcaga ccaaaacgat ctcaagaaga
     6541 tcatcttatt aatcagataa aatatttcta gatttcagtg caatttatct cttcaaatgt
     6601 agcacctgaa gtcagcccca tacgatataa gttgtaattc tcatgttagt catgccccgc
     6661 gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt
     6721 gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg
     6781 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     6841 cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc
     6901 cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag
     6961 gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc
     7021 gtcgtatccc actaccgaga tgtccgcacc aacgcgcagc ccggactcgg taatggcgcg
     7081 cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc
     7141 attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc
     7201 cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg
     7261 cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac
     7321 cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg
     7381 tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc
     7441 aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag
     7501 aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac
     7561 cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc
     7621 gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg
     7681 ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg
     7741 cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac
     7801 accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg
     7861 actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc
     7921 cgggatctcg acgctctccc ttatgcgact cctgcattag gaaattaata cgactcacta
     7981 ta
//
