LOCUS       sequence-463645-        6339 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CageIL-2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6339)
  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 6339)
  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-0013 CMVTO-SumIL2-rens-IL2Rb
FEATURES             Location/Qualifiers
     source          1..6339
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(570..1361,1362..1430))
                     /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     1193..1212
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1431..1535)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1610..1629)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1796..2175
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2176..2379
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2330..2350
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    2381..2399
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2402..2420
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2430..2454
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             3733..3762
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(3818..3835)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3824..4048
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4220..4239)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4371..4418
                     /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             4426..5448
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4426..5448
                     /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    5578..5699
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5615..5634)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5669..5688
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5748..5764)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5748..5764)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5761..5783)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5772..5788
                     /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(5796..5802,5803..5820,5821..5826))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5841..5862
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5979..5996)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(6150..6339,1..399))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6230..6249)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
       61 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
      121 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
      181 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
      241 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
      301 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
      361 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
      421 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
      481 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
      541 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
      601 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
      661 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
      721 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
      781 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
      841 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
      901 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
      961 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     1021 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     1081 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     1141 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     1201 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     1261 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     1321 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     1381 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     1441 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     1501 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     1561 gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt acaatctgct
     1621 ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt
     1681 agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga
     1741 atctgcttag ggttaggcgt tttgcgctgc ttcgtacggg ccagatatac gcgttgacat
     1801 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
     1861 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
     1921 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
     1981 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     2041 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     2101 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
     2161 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
     2221 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
     2281 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
     2341 ggtaggcgtg tacggtggga ggtctatata agcagagctc tccctatcag tgatagagat
     2401 ctccctatca gtgatagaga tcgtcgacga gctcgtttag tgaaccgtca gatcgcctgg
     2461 agacgccatc cacgctgttt tgacctccat agaagacacc gggaccgatc cagcctccgg
     2521 actctagcgt ttaaacttaa gcttcgccac catgtacagg atgcaactcc tgtcttgcat
     2581 tgcactaagt cttgcacttg tcacaaacgc tagcgctcca acctccagct ctacaaaaaa
     2641 gactcagctg cagctggagc acctgctgct cgacctacaa atgatactca acggaatcaa
     2701 caactataag aatcctaagc tgacgcggat gctgaccgcc aagttttaca tgcccaaaaa
     2761 ggcgaccgag ctgaagcatt tgcagtgtct ggaggaagag ctcaagcctt tggaggaggt
     2821 gcttaatctc gcccaaagca aaaatttcca ctttgatccg cgggacgtcg tgtcaaatat
     2881 caacgtcttt gtgctggaac tgaaaggctc tgagacaacc tttatgtgtg aatatgccga
     2941 tgaaactgct accatcgttg agtttttaaa tcgatggata acattttgcc agagtataat
     3001 ttctactttg accggcgccg ggacaggata tattcaccca ttccatctgg ttattcataa
     3061 cgagtcgggc ggtgggggat ccgcggtgaa tggcacttcc cagttcacat gcttctacaa
     3121 ttcgagagcc aacatctcct gtgtctggag ccaagatggg gctctgcagg acacttcctg
     3181 ccaagtccat gcctggccgg acagacggcg gtggaaccaa acctgtgagc tgctccccgt
     3241 gagtcaagca tcctgggcct gcaacctgat cctcggagcc ccagattctc agaaactgac
     3301 cacagttgac atcgtcaccc tgagggtgct gtgccgtgag ggggtgcgat ggagggtgat
     3361 ggccatccag gacttcaagc cctttgagaa ccttcgcctg atggccccca tctccctcca
     3421 agttgtccac gtggagaccc acagatgcaa cataagctgg gaaatctccc aagcctccca
     3481 ctactttgaa agacacctgg agttcgaggc ccggacgctg tccccaggcc acacctggga
     3541 ggaggccccc ctgctgactc tcaagcagaa gcaggaatgg atctgcctgg agacgctcac
     3601 cccagacacc cagtatgagt ttcaggtgcg ggtcaagcct ctgcaaggcg agttcacgac
     3661 ctggagcccc tggagccagc ccctggcctt caggacaaag cctgcagccc ttgggaagga
     3721 cacctagggc ccgaacaaaa actcatctca gaagaggatc tgaatagcgc cgtcgaccat
     3781 catcataatc atcattgagt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     3841 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     3901 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     3961 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     4021 catgctgggg atgcggtggg ctctatggag atctatcgat gttctgggga tgcggtgggc
     4081 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     4141 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     4201 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     4261 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     4321 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtaccta gaagttccta
     4381 ttccgaagtt cctattctct agaaagtata ggaacttcct tggccaaaaa gcctgaactc
     4441 accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg
     4501 cagctctcgg agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat
     4561 gtcctgcggg taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac
     4621 tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca ttggggaatt cagcgagagc
     4681 ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc
     4741 gaactgcccg ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat
     4801 cttagccaga cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca
     4861 tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg
     4921 gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag
     4981 gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg
     5041 gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa
     5101 tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg
     5161 cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg
     5221 ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca
     5281 gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt
     5341 acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc
     5401 gatagtggaa accgacgccc cagcactcgt ccgagggcaa aggaatagca cgtactacga
     5461 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     5521 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     5581 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     5641 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     5701 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     5761 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     5821 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     5881 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     5941 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     6001 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     6061 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     6121 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     6181 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     6241 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     6301 tacctgtccg cctttctccc ttcgggaagc gtggcgctt
//
