LOCUS       sequence-358714-        7388 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses CuCBP and BaSP in E. coli BL21(DE3).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7388)
  AUTHORS   Schwaiger KN, Voit A, Wiltschi B, Nidetzky B
  TITLE     Engineering cascade biocatalysis in whole cells for bottom-up 
            synthesis of cello-oligosaccharides: flux control over three 
            enzymatic steps enables soluble production.
  JOURNAL   Microb Cell Fact. 2022 Apr 9;21(1):61. doi: 
            10.1186/s12934-022-01781-w.
  PUBMED    35397553
REFERENCE   2  (bases 1 to 7388)
  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"; journalName: "Microb Cell
            Fact."; date: "2022-04-9"; volume: "21(1):61. doi"; pages: " 
            10.1186/s12934-022-01781-w"
COMMENT     Sequence Label: pBICI_high ori
FEATURES             Location/Qualifiers
     source          1..7388
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     10..29
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        10..28
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    29..53
                     /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             69..89
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             109..126
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             2608..2619
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2635..2658
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     terminator      complement(4186..4233)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      4240..4334
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      4367..4955
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4856..4875
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5109..5126
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5141..5281
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5207..5226)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(join(5341..6132,6133..6201))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5964..5983
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     RBS             6208..6219
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     primer_bind     7272..7291
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7311..7388)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
ORIGIN
        1 gaattcagtt aatacgactc actatagggg aattgtgagc ggataacaat tcccctctag
       61 aaataatgtt gtttaacttt aagaaggaga tatcatatgg gcagcagcca tcaccatcac
      121 catcacgggt cgacccgcta tggtcatttt gacgatgaag cacgcgagta cgttatcacc
      181 acaccgcata ctccgtaccc ttggatcaac tatctgggtt cggaacagtt cttcagtctt
      241 ttgtcgcacc aggctggcgg ctattcgttt tatcgcgatg ccaagatgcg tagattgaca
      301 agatacagat acaacaacat accagcagac gcgggaggcc ggtatcttta tgtaaacgac
      361 ggtggggacg tttggacgcc atcgtggctt cctgttaaag cagacttaga tcattttgaa
      421 gccagacatg gtcttggcta cagtaccatt acaggtgaaa gaaatggggt acgggttgag
      481 accttgtttt tcgtcccagt cggtgagaac gcggaggtcc agaaagttac ggtcactaat
      541 acctctgact cgtataaatc cctgacgttg ttttcttttg tcgaattttg tctttggaac
      601 gcgcaagacg atcaaactaa ctaccagcgt aatctgagca taggagaggt tgaagtggaa
      661 caagaatctc cgcatggcag tgcaatttat caccgtacag agtaccggga gagacgcgat
      721 cactacgcgg tattcgccgt taatacgcag gccgagggtt tcgatacaga tcgcgacaca
      781 tttgtgggag cctataactc gcttggtgaa gccgcagttc cccttaaggg agaaagtgca
      841 aatagcgttg caagtggatg gtatcctatt ggctctcata gcgtggcagt gtctttagcg
      901 cctggagaat ccagagaatt ggtttatgtg cttgggtacg tggaaaatcc tgatgaggag
      961 aagtgggcag acgacgcaaa acaggtcgtg aataaagaaa gagctcacgc tttattatca
     1021 agatttgcta cgtccgaaca gactgacgca gctttcgccg ctttgaagga ttattggacg
     1081 gatttgttat cgacgtattc cgttagttcc aatgacgaaa aattggaccg gatggttaat
     1141 atatggaatc agtaccaatg catggtaacc tttaacatga gtcgctcggc gagctttttc
     1201 gagaccggta ttgggagagg gatggggttt cgtgatagca accaagacct tctgggcttt
     1261 gttcacttaa tcccagagcg ggcgcgtgag cgcatcattg acatagcgtc aacccagttc
     1321 gcggacggtt ccgcttatca ccaataccaa cctttaacga agcgcggtaa caacgacatt
     1381 ggctctgggt ttaacgacga tcctctttgg ttaatcgcgg gaactgccgc ctacatcaaa
     1441 gaaactggtg acttctccat attagatgag cccgtacctt ttgataacga accggggtca
     1501 gaagttccgt tatttgagca cctgacccgt agttttgagt ttaccgtaac ccatcggggt
     1561 ccccacggtt tgcctcttat tggccgcgca gattggaacg attgtttgaa tcttaattgc
     1621 ttcagtacaa ccccgggcga gtcatttcag acaacggaga accaagccgg tggcgtagcg
     1681 gagtcaacgt tcatagccgc gcaattcgtt ttatatgggg agcagtatgc cgagttggca
     1741 gcgcggcgcg ggttggccga cgtagccgat agagcaagag ggcatgtggc tgaaatgaga
     1801 gacgcacttc tgaccgacgg gtgggacggc agctggttct tgagagctta tgactattat
     1861 ggaaacccta tcggcaccga cgcacatgac gaagggaaga tttggatcga accgcaaggg
     1921 ttcgcggtga tggctggagt cggtgtaggc gaagggcctc aagatacgga cgctcccgct
     1981 attaaggcgc ttgattctgt gaacgaaatg ttggcaaccg atcacggtat ggtccttcaa
     2041 tacccagcat acactacata ccaagtacac atgggtgaag tatccactta tccacccggg
     2101 tataaagaga atggaggtat attttgccac aataatcctt gggttatcat agcggaaact
     2161 gtcgtcggaa gaggcggacg ggcatttgat tactacaaga gaattactcc cgcgtatcgc
     2221 gaggacataa gcgacgttca ccgtcttgaa ccctatgtct atgcacaaat gatagcaggg
     2281 aaggaggcgg tccgtcacgg tgaagcaaag aacagctggc ttaccgggac tgcggcttgg
     2341 aactttgtga ccgtgtcaca gtaccttctt ggggttcgcc cagagtacga cggcttggtg
     2401 gtagacccgc agatcggacc tgacgtacct agctttacgg ttacgcgtgt cgcccgtgga
     2461 gccacttacg agataacagt cactaacagt gggaccgacg gatcacgcgg tcgccttgtc
     2521 gttgatggaa cacctgtaga aggcaacttg gttccttatg cccccgcagg gtcaacagtt
     2581 cgtgtcgatg tgacattgta gtgtacaaaa gaggagaaaa cccccatggc aagctggagc
     2641 cacccgcagt tcgaaaaggg taccatgaaa aataaagtcc aactgattac ctatgcggat
     2701 cgtctgggtg atggcaccat taaaagcatg acggacatcc tgcgtacccg ctttgatggc
     2761 gtttatgacg gtgtccacat tctgccgttt ttcaccccgt ttgatggcgc cgacgcaggt
     2821 ttcgatccga tcgaccatac caaagttgat gaacgtctgg gtagctggga tgacgtcgcc
     2881 gaactgtcta aaacccacaa cattatggtg gatgcaatcg ttaatcacat gtcatgggaa
     2941 tcgaaacagt tccaagatgt tctggccaaa ggcgaagaat cagaatatta cccgatgttt
     3001 ctgaccatga gcagcgtgtt cccgaacggt gccacggaag aagacctggc aggcatttat
     3061 cgtccgcgtc cgggtctgcc gtttacccac tacaaattcg caggcaaaac gcgtctggtc
     3121 tgggtgtctt ttaccccgca gcaagtggac atcgatacgg acagtgataa aggttgggaa
     3181 tatctgatgt ccattttcga ccaaatggcg gccagccatg tgtcttatat ccgcctggat
     3241 gcggttggct acggtgccaa agaagcaggc accagttgct ttatgacccc gaaaacgttc
     3301 aaactgattt cccgtctgcg cgaagaaggc gtgaaacgcg gtctggaaat tctgatcgaa
     3361 gtccacagct attacaaaaa acaggtggaa atcgcgtcta aagtggatcg cgtttatgac
     3421 tttgccctgc cgccgctgct gctgcatgcc ctgagtaccg gccacgtcga accggtggcc
     3481 cattggacgg atattcgtcc gaacaatgct gttaccgtcc tggatacgca tgacggcatc
     3541 ggcgttattg atatcggttc agatcagctg gaccgctcgc tgaaaggtct ggtgccggac
     3601 gaagatgtgg ataacctggt gaatacgatt catgcgaaca cccacggcga atcacaggca
     3661 gctaccggtg cggcggcatc gaacctggat ctgtaccaag ttaatagtac ctactactcc
     3721 gctctgggct gtaacgatca gcactacatc gctgcgcgtg cggtgcagtt tttcctgccg
     3781 ggtgttccgc aagtctatta cgtgggcgcc ctggcaggta aaaatgatat ggaactgctg
     3841 cgcaaaacca acaatggtcg tgacattaac cgccattatt acagcacggc cgaaatcgat
     3901 gaaaatctga aacgtccggt ggttaaagca ctgaacgctc tggcgaaatt tcgcaatgaa
     3961 ctggatgctt ttgacggcac cttctcatat accacggatg acgatacgag tatttccttt
     4021 acctggcgtg gtgaaacgtc gcaggccacc ctgacgttcg aaccgaaacg cggcctgggt
     4081 gttgataata ccacgccggt cgcaatgctg gaatgggaag acagtgctgg tgaccatcgc
     4141 tccgacgacc tgatcgctaa tccgccggtt gttgcgtaaa agcttcaaaa aacccctcaa
     4201 gacccgttta gaggccccaa ggggttatgc taggagctcg actcctgttg atagatccag
     4261 taatgacctc agaactccat ctggatttgt tcagaacgct cggttgccgc cgggcgtttt
     4321 ttattggtga gaatggatcc cccgtagaaa agatcaaagg atcttcttga gatccttttt
     4381 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     4441 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     4501 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     4561 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     4621 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     4681 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     4741 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     4801 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     4861 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     4921 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     4981 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     5041 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     5101 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc
     5161 ttacgcatct gtgcggtatt tcacaccgca tatggtgcac tctcagtaca atctgctctg
     5221 atgccgcata gttaagccag tatacactcc gctatcgcta cgtgactggg tcatggctgc
     5281 gccccgacac ccgccaacct cgagtaaaaa aaatccttag ctttcgctaa ggatctgcag
     5341 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     5401 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     5461 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     5521 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     5581 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     5641 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     5701 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     5761 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     5821 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     5881 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     5941 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     6001 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     6061 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     6121 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     6181 acggaaatgt tgaatactca tgcttaattt ctcctcttta atagatcttc actgcccgct
     6241 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     6301 ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag
     6361 ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg
     6421 ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc
     6481 ctcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc cggactcggt
     6541 aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac
     6601 gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc
     6661 ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag
     6721 acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc
     6781 caatgcgacc agatgctcca cgcccagtcg cgtaccgtcc tcatgggaga aaataatact
     6841 gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc
     6901 ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg
     6961 ttgcgcgaga agattgtgca ccgcccttta caggcttcga cgccgcttcg ttctaccatc
     7021 gacaccacca cgctggcacc cagttgatcg gcgcgagatt taatcgccgc gacaatttgc
     7081 gacggcgcgt gcagggccag actggaggtg gcaacgccaa tcagcaacga ctgtttgccc
     7141 gccagttgtt gtgccacgcg gttgggaatg taattcagct ccgccatcgc cgcttccact
     7201 ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga aacggtctga
     7261 taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac attcaccacc
     7321 ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt gcaccattcg
     7381 atggtgtc
//
