LOCUS       sequence-469300-        8603 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of Cingulin (human) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8603)
  AUTHORS   Vasileva E, Spadaro D, Rouaud F, King JM, Flinois A, Shah J, 
            Sluysmans S, Mean I, Jond L, Turner JR, Citi S
  TITLE     Cingulin binds to the ZU5 domain of scaffolding protein ZO-1 to 
            promote its extended conformation, stabilization, and tight junction
            accumulation.
  JOURNAL   J Biol Chem. 2022 Apr;298(4):101797. doi: 10.1016/j.jbc.2022.101797.
            Epub 2022 Mar 5.
  PUBMED    35259394
REFERENCE   2  (bases 1 to 8603)
  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: "J Biol 
            Chem."; date: "2022-04"; volume: "298(4):101797. doi"; pages: " 
            10.1016/j.jbc.2022.101797. Epub 2022 Mar 5"
COMMENT     Sequence Label: hCGN-HA
FEATURES             Location/Qualifiers
     source          1..8603
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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)."
     primer_bind     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    1040..1161
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1051..1070)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1105..1124
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1291..1665)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     promoter        complement(1684..1731)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(1779..2108)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1793..1928
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1847..1866)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     2093..2113
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(2122..2550)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2232..2253)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2444..2463
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    2596..2820
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2809..2826
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(2855..2881)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     RBS             5106..5114
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     regulatory      6493..6502
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(6555..6574)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6556..6574)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(6619..6822)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(6648..6668)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        6823..7202
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     7374..7393
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        7468..7572
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7573..7641,7642..8433)
                     /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     complement(7791..7810)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagctct agctagaggt
     1021 cgacggtata cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg
     1081 cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt
     1141 ataagctgca ataaacaagt tggggtgggc gaagaactcc agcatgagat ccccgcgctg
     1201 gaggatcatc cagccggcgt cccggaaaac gattccgaag cccaaccttt catagaaggc
     1261 ggcggtggaa tcgaaatctc gtagcacgtg tcagtcctgc tcctcggcca cgaagtgcac
     1321 gcagttgccg gccgggtcgc gcagggcgaa ctcccgcccc cacggctgct cgccgatctc
     1381 ggtcatggcc ggcccggagg cgtcccggaa gttcgtggac acgacctccg accactcggc
     1441 gtacagctcg tccaggccgc gcacccacac ccaggccagg gtgttgtccg gcaccacctg
     1501 gtcctggacc gcgctgatga acagggtcac gtcgtcccgg accacaccgg cgaagtcgtc
     1561 ctccacgaag tcccgggaga acccgagccg gtcggtccag aactcgaccg ctccggcgac
     1621 gtcgcgcgcg gtgagcaccg gaacggcact ggtcaacttg gccatggttt agttcctcac
     1681 cttgtcgtat tatactatgc cgatatacta tgccgatgat taattgtcaa cacgtgctga
     1741 tcagatccga aaatggatat acaagctccc gggagctttt tgcaaaagcc taggcctcca
     1801 aaaaagcctc ctcactactt ctggaatagc tcagaggcag aggcggcctc ggcctctgca
     1861 taaataaaaa aaattagtca gccatggggc ggagaatggg cggaactggg cggagttagg
     1921 ggcgggatgg gcggagttag gggcgggact atggttgctg actaattgag atgcatgctt
     1981 tgcatacttc tgcctgctgg ggagcctggg gactttccac acctggttgc tgactaattg
     2041 agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc acaccctaac
     2101 tgacacacat tccacagaat taattcgcgt taaatttttg ttaaatcagc tcatttttta
     2161 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     2221 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     2281 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     2341 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     2401 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     2461 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     2521 ccgcgcttaa tgcgccgcta cagggcgcgt ggggataccc cctagagccc cagctggttc
     2581 tttccgcctc agaagccata gagcccaccg catccccagc atgcctgcta ttgtcttccc
     2641 aatcctcccc cttgctgtcc tgccccaccc caccccccag aatagaatga cacctactca
     2701 gacaatgcga tgcaatttcc tcattttatt aggaaaggac agtgggagtg gcaccttcca
     2761 gggtcaagga aggcacgggg gaggggcaaa caacagatgg ctggcaacta gaaggcacag
     2821 tcgaggctga tcagcgggtt taaacgggcc cctaagcgta gtctgggacg tcgtatgggt
     2881 atctagaaca ggagctggtc tgtaggttgc tctccgtaag cagtgatgca atggacgagg
     2941 gatcgtagac actgtcgaat tcctcatctg agctcagccc ttcgtttttg agagctgact
     3001 cagcagctga gcgggaagct ttgcgccagg agtccttctc cagagacttg atccgggcct
     3061 ggagctgttc attgacctca tgctgctcct ccacctcacg ctgggccttc ttcctcaggc
     3121 cgtccagtcg ctcaatttcc tcttctgctt catccacctg acgcttcaaa gccttcaccc
     3181 tcaggcttag ctggtctttc tggtcattga catgctgccg ctcgtcttca atctggatgg
     3241 atagttcttt aactttccgc tccagttttc gattggtaga ctgcagaact gtcttctccc
     3301 tctcttcagc ctgtagccgc tcctgcaaca actgattctg ggactcaagc tgagagaggc
     3361 tggcactagg cttctggaag ccttctgagc tggccaaccg ggtcttcagg tccttgttct
     3421 gtctctccaa ggagattttg tcacactcca ggtcctgccg agcagacctt tcctgcatga
     3481 gctctgtcct cagctgatcc acctggtccc ggccacgatt cacccgatct gttagcagct
     3541 ccacggtgtt cttctcctca tctaactctg tttccagccg tgagactttt tcctcgagac
     3601 ccttcagctg ccgggccctg tcgtcctggg aacgcttctt gttctctgcc tcttgctcca
     3661 gcccctgcag tcgctgggcc agtagctctt tgtccagccg ggctgtgtcc cgctcagcct
     3721 gggatgcctg cagggcctgc cgcagcctct ggatctcatc ctgaagtcgg ctcagtcctc
     3781 cagaggtctt ctcagcctca ctggcccaat ccttggcctg gcgctgggca tctgccacct
     3841 cccgccgggc cttttcctta taatcctcca gctgggcctg gagctgctgc agggcttgct
     3901 tagagtcctc cccgatcttc tccaactcct tgttcagtcg gtccacagtc ctgtccagca
     3961 aacgcttctg ctcctccagc tcagccttgc ctcgccggag cacctcccgc tgcttccctt
     4021 cctcctccag ggcccggttc agtgtctgct gctcctgccc caggcgggcc agcccccgct
     4081 gagcctcctc taggcgtgcc tccagtgccc gcttggctgc tgccagactc ccctcctctt
     4141 cctgggacgc attcagggcc tcctccagct gctgtttctc tgcctccagc cgctgcagct
     4201 tgtcccgtag tcgtgcctcc accgcttccc caccatccac cagacctcga gtctccttca
     4261 gctgctgctc caaacccagg atgcgccggc ggaattcgtc attttcctcc tgggtctccc
     4321 gaagcgtcgt ctccactgcg gcccgccgct gccccagcac tgttgcctct gcctcggcca
     4381 ccatcttagc cttggaagcc tcttcacagt cctgtcgcag ttgctggagg gtcttttgta
     4441 gctggaggtt ctgttcttcc agctcccgac ctcgatcagc ctcgaccctc aggaccgcca
     4501 gctgcttctc caactcccgg tcccggtgtc gcccagccac ctcctggctc tgccgttctg
     4561 cctgcagttc cttaagctcc tcctgtgtcc ggagcagctc cttcttgagc acctcaacct
     4621 ggcgagtatc tccagcacta gctcgggcct gctctaattc cctctgcaag acctctatct
     4681 tctctccaag ctcctcttcc atctcctcct tctccattcg cagctgcagg agttcctgct
     4741 tggtggctct aagatcctcc ttgttcttct ggaacatact ctgccaatgc cctgtctcct
     4801 ctgaagtctc ctccagctcc ctctgcagcc ctcgcacaag ggctgacatc ttctgcctct
     4861 cggcctccag cactgcatgg tcctgggttg catcttgcat gctcctgcgg agctgctctg
     4921 tgtctcgctg gtactgctgc cggacatgtt ccacctcctg gtcacgggag gctacctcct
     4981 ctttcagggc ccccttcagg gctgtcaact cccgctcccg cagcctcagc tgctcctcta
     5041 ctcgctgttt cccctccaag acctcttcca gaagttcccg ggtctctaac aggtccttca
     5101 gtaacacctc cttagcaggc tcaggaccct ggacatgttt cagcttgttc tgcagctcca
     5161 tcacctgggt ctccagccca tgtcgtaaat cttcaccctg gtccaagagg cgcttcatgt
     5221 tctggagctc cttgttgctc tgctgggcct cccccttcct cctctccagc agctcctgca
     5281 gtcggctgca ctcttctgtt ttctcctcca gctgccgctc cagcccaact tgggatggct
     5341 ctagcttctg ccgcttcttc acctcttcat ccagctttcg ctgtagctcc tccacttttc
     5401 gggtaagctc accctgccct gccacggcct tagtagaacc agaagaaacc atcactagag
     5461 gctgcatctt ctccagcacc aaactaacct tcctcctcac agaggtttca ctttctgagc
     5521 ttccctccct caggatgcca tagatggtgg ccttcatatg gtccacactg cctggtgggg
     5581 ctgcctcctg ctggtctcga aggaggtctg gagttgattt gaactgcagc atggtggggt
     5641 cctgtgcact cctccgcggc tcctcaaaac tctgaaggac ccagtcctga gtctgacgag
     5701 aacggctaaa gccactgaga ggactcaggt tctggctctg ggctggctgc ttcgaagtgc
     5761 ccacatggtt gtcatatttt gtgctagagg tccagtggtt ggtggactgg cgctcccgtt
     5821 cctcaaaggt gtcccgtggg aggcggctgt ccaggctctt gctccgtttg cgctgttcag
     5881 ggggtagcat ccgtgttcgg cggccagtcc gaccccgggc ctggcctcca agttgactgt
     5941 caaacttgtt gatgagtgag tccactgaag acaggggagc agtgtcaatg gtgctacctg
     6001 gggaagccac tttcggggct agctccagca tgctgttgct tctgttacta gggtccactg
     6061 ggccagggcc tgccagtgag gcctgggagt gggaacggag tagctttcca ttccagtagg
     6121 ccccaggctc ctcatcagat gtgctgctct gcgagggggc aggaaatccc ttgacctgag
     6181 agtaggggtt ctcagggagt tccaaatctg agctcagagc tcctgaggcc ccttggtcat
     6241 tggccccctt gatttggacc ccaaaggagt caccgccttt ctccccactg ttgagcacca
     6301 caaagggctg cccagcgatt ccctgcacac gcacagcaac cccgtaggta ctggctcttg
     6361 catccttagc tgggcgtcgt ccacctcgac gtagagtgcc catctctgca ccactcactg
     6421 gctctgtgat gaagcgaatc tggactccat ggtctacggg gccccggggc tcagccatgt
     6481 tgggtgcctg ctccatggtg gcggatccga gctcggtacc aagcttaagt ttaaacgcta
     6541 gccagcttgg gtctccctat agtgagtcgt attaatttcg ataagccagt aagcagtggg
     6601 ttctctagtt agccagagag ctctgcttat atagacctcc caccgtacac gcctaccgcc
     6661 catttgcgtc aatggggcgg agttgttacg acattttgga aagtcccgtt gattttggtg
     6721 ccaaaacaaa ctcccattga cgtcaatggg gtggagactt ggaaatcccc gtgagtcaaa
     6781 ccgctatcca cgcccattga tgtactgcca aaaccgcatc accatggtaa tagcgatgac
     6841 taatacgtag atgtactgcc aagtaggaaa gtcccataag gtcatgtact gggcataatg
     6901 ccaggcgggc catttaccgt cattgacgtc aatagggggc gtacttggca tatgatacac
     6961 ttgatgtact gccaagtggg cagtttaccg taaatactcc acccattgac gtcaatggaa
     7021 agtccctatt ggcgttacta tgggaacata cgtcattatt gacgtcaatg ggcgggggtc
     7081 gttgggcggt cagccaggcg ggccatttac cgtaagttat gtaacgcgga actccatata
     7141 tgggctatga actaatgacc ccgtaattga ttactattaa taactagtca ataatcaatg
     7201 tcaacgcgta tatctggccc gtacatcgcg aagcagcgca aaacgcctaa ccctaagcag
     7261 attcttcatg caattgtcgg tcaagccttg ccttgttgta gcttaaattt tgctcgcgca
     7321 ctactcagcg acctccaaca cacaagcagg gagcagatac tggcttaact atgcggcatc
     7381 agagcagatt gtactgagag tgcaccatag gggatcggga gatctcccga tccgtcgacg
     7441 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     7501 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     7561 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     7621 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     7681 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     7741 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     7801 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     7861 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     7921 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     7981 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     8041 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     8101 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     8161 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     8221 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     8281 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     8341 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     8401 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     8461 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     8521 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     8581 gtagaaaaga tcaaaggatc ttc
//
