LOCUS       sequence-459380-        6915 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  C-CageIL-6.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6915)
  AUTHORS   Aldrete CA, Call CC, Sant'Anna LE, Vlahos AE, Pei J, Cong Q, Gao XJ
  TITLE     Orthogonalized human protease control of secreted signals.
  JOURNAL   Nat Chem Biol. 2025 Jan 15. doi: 10.1038/s41589-024-01831-x.
  PUBMED    39814991
REFERENCE   2  (bases 1 to 6915)
  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: "Nat Chem 
            Biol. 2025 Jan 15. doi: 10.1038/s41589-024-01831-x."
COMMENT     Sequence Label: CA-0034 CMVTO-hIL6-rens-hIL6Ra_M4
FEATURES             Location/Qualifiers
     source          1..6915
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(403..1194,1195..1263))
                     /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     1026..1045
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1264..1368)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1443..1462)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1629..2008
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2009..2212
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2163..2183
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    2214..2232
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2235..2253
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2263..2287
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4142..4171
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(4227..4244)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4233..4457
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4629..4648)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4780..4827
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             4835..5857
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4835..5857
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    5987..6108
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6024..6043)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6078..6097
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6157..6173)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6157..6173)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6170..6192)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6181..6197
                     /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)."
     promoter        complement(join(6205..6211,6212..6229,6230..6235))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6250..6271
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6388..6405)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(6559..6915,1..232))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6639..6658)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
       61 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
      121 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
      181 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
      241 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
      301 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
      361 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
      421 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
      481 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
      541 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
      601 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
      661 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
      721 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
      781 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
      841 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
      901 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
      961 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     1021 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     1081 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     1141 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     1201 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     1261 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     1321 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     1381 aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc
     1441 agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg
     1501 gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac
     1561 aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgtac gggccagata
     1621 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
     1681 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
     1741 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
     1801 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     1861 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
     1921 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
     1981 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
     2041 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
     2101 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
     2161 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctccctat
     2221 cagtgataga gatctcccta tcagtgatag agatcgtcga cgagctcgtt tagtgaaccg
     2281 tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagac accgggaccg
     2341 atccagcctc cggactctag cgtttaaact taagcttcgc caccatgaac agcttctcta
     2401 ccagcgcttt tggacccgtg gccttttcgt tgggcctgct tctcgttctt cctgccgcat
     2461 tccctgcacc agtcccccca ggcgaagatt ctaaagatgt ggctgctcca cacagacagc
     2521 cccttactag cagtgaacgt attgataaac agatcaggta catcctggac ggaatctcag
     2581 ccctaagaaa agagacatgc aataagtcca acatgtgtga gagttccaag gaggccctcg
     2641 ccgagaataa cctgaatcta cccaagatgg ccgagaagga cggttgtttc cagagtgggt
     2701 ttaacgaaga gacttgcctg gtgaaaatta taaccgggct gctggaattc gaagtgtacc
     2761 tggagtattt gcaaaatcgg ttcgagtcat ctgaagagca ggccagggct gtccagatgt
     2821 ctaccaaagt attaatccag ttcttgcaga agaaagctaa gaacctggat gcaataacaa
     2881 ctcctgaccc caccacaaac gcgagcctgc ttacaaagct ccaggcgcag aatcagtggt
     2941 tacaagacat gacgacccat ctgattctcc gatcatttaa ggaatttttg caatcctccc
     3001 tccgggcact gcgccaaatg accggtggcg gcggtagcgg atatattcac ccattccatc
     3061 tggttattca taacgagtcg ggcggtggta ccggccttgc gccgagaaga tgccccgccc
     3121 aagaggtggc acggggagtt ctgaccagtc tcccaggcga ttctgtgact ctgacctgcc
     3181 ctggtgtaga accggaagac aatgcaaccg tccactgggt gttgagaaag ccagcagcgg
     3241 gaagccatcc ctccaggtgg gccggcatgg gacgacgcct gctgctgagg agtgtgcagc
     3301 tacacgattc tggtaactac tcttgttata gggccggccg gccggctggc accgtccatt
     3361 tactcgtgga tgtgcctccc gaggagcccc aattgagttg ctttcgaaaa agcccactgt
     3421 ccaatgtagt ctgcgagtgg ggtccacggt cgaccccctc cctcacaacg aaagcggtct
     3481 tacttgttag gaagttccag aacagccctg ctgaggactt ccaggaacct tgtcagtaca
     3541 gtcaagagtc tcagaagttt agctgtcaat tggcagtccc agagggggac agcagctttt
     3601 atatcgtatc aatgtgcgtg gcctcctccg tggggagtaa gttctccaag acccagactt
     3661 tccagggctg tggcattctt cagcctgacc caccagccaa cattactgtg accgccgtag
     3721 acagagatcc ccgttggctg agcgttactt ggcaagaccc ccatagttgg aactcatctt
     3781 tctatcggct ccgctttaag ctccgttacc gcgccgaacg cagcaagaca tttacgacat
     3841 ggatggtcaa agatcttcag caccactgtg tgatatctgt ggcttggtcc gggctgaggc
     3901 acgttgtcca gctgagagcc caggaggaat ttggacaggg ggagtggtcc gagtggtcac
     3961 cagaagctat ggggactcct tggacagaat ctcgatcccc ccccgctgag aatgaagtgt
     4021 caacccccat gcaggcactg acaacaaata aagatgacga taatatccta ttccgggact
     4081 cagccaacgc tacttcgttg cctgttcagg acagctcttc agtgcctctg ccttagggcc
     4141 cgaacaaaaa ctcatctcag aagaggatct gaatagcgcc gtcgaccatc atcataatca
     4201 tcattgagtt taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     4261 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     4321 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     4381 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     4441 tgcggtgggc tctatggaga tctatcgatg ttctggggat gcggtgggct ctatggcttc
     4501 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     4561 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     4621 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     4681 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     4741 ccccaaaaaa cttgattagg gtgatggttc acgtacctag aagttcctat tccgaagttc
     4801 ctattctcta gaaagtatag gaacttcctt ggccaaaaag cctgaactca ccgcgacgtc
     4861 tgtcgagaag tttctgatcg aaaagttcga cagcgtctcc gacctgatgc agctctcgga
     4921 gggcgaagaa tctcgtgctt tcagcttcga tgtaggaggg cgtggatatg tcctgcgggt
     4981 aaatagctgc gccgatggtt tctacaaaga tcgttatgtt tatcggcact ttgcatcggc
     5041 cgcgctcccg attccggaag tgcttgacat tggggaattc agcgagagcc tgacctattg
     5101 catctcccgc cgtgcacagg gtgtcacgtt gcaagacctg cctgaaaccg aactgcccgc
     5161 tgttctgcag ccggtcgcgg aggccatgga tgcgatcgct gcggccgatc ttagccagac
     5221 gagcgggttc ggcccattcg gaccgcaagg aatcggtcaa tacactacat ggcgtgattt
     5281 catatgcgcg attgctgatc cccatgtgta tcactggcaa actgtgatgg acgacaccgt
     5341 cagtgcgtcc gtcgcgcagg ctctcgatga gctgatgctt tgggccgagg actgccccga
     5401 agtccggcac ctcgtgcacg cggatttcgg ctccaacaat gtcctgacgg acaatggccg
     5461 cataacagcg gtcattgact ggagcgaggc gatgttcggg gattcccaat acgaggtcgc
     5521 caacatcttc ttctggaggc cgtggttggc ttgtatggag cagcagacgc gctacttcga
     5581 gcggaggcat ccggagcttg caggatcgcc gcggctccgg gcgtatatgc tccgcattgg
     5641 tcttgaccaa ctctatcaga gcttggttga cggcaatttc gatgatgcag cttgggcgca
     5701 gggtcgatgc gacgcaatcg tccgatccgg agccgggact gtcgggcgta cacaaatcgc
     5761 ccgcagaagc gcggccgtct ggaccgatgg ctgtgtagaa gtactcgccg atagtggaaa
     5821 ccgacgcccc agcactcgtc cgagggcaaa ggaatagcac gtactacgag atttcgattc
     5881 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     5941 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     6001 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     6061 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     6121 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     6181 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     6241 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     6301 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6361 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     6421 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     6481 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     6541 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     6601 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     6661 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     6721 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     6781 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     6841 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     6901 actggcagca gccac
//
