LOCUS       sequence-459372-        5745 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CleaveIL-10.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5745)
  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 5745)
  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-0165 CMVTO-mono-hIL10 (R5A11M)-rens
FEATURES             Location/Qualifiers
     source          1..5745
                     /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             2972..3001
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(3057..3074)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3063..3287
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3459..3478)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3610..3657
                     /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             3665..4687
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3665..4687
                     /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    4817..4938
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4854..4873)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4908..4927
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4987..5003)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4987..5003)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5000..5022)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5011..5027
                     /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(5035..5041,5042..5059,5060..5065))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5080..5101
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5218..5235)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(5389..5745,1..232))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5469..5488)
                     /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 caccatgcac tcctccgcgc
     2401 tcttgtgttg cctagtttta ttgaccggag tccgcgcatc acctgggcaa ggcactcaga
     2461 gtgaaaattc ttgcactcac ttccctggaa ttctccccaa catgctcaga gacctgcgag
     2521 atgctttctc tagagtcaaa acgttctttc aaatgaagga ccagctagat aatcttttgc
     2581 tgaaagagtc ccttctcgag gattttaaag gctatctggg ctgtcaggct cttagcgaga
     2641 tgatacagtt ctacctcgag gaggtgatgc cacaggctga gaaccaagac ccagacatta
     2701 aagcccatgt aatctcactg ggggaaaacc tgaacacatt acgcctgcgg ctgcggaggt
     2761 gccataggct gctgccttgt gaaaattata ttcacccatt ccatctggtt attcataacg
     2821 agtcgaagag caaggccgtg gaacaggtga agaatgcctt taataagctg caggaaaagg
     2881 ggatctacaa ggcaatgagc gagtttgata tattcatcaa ctatatcgag gcctacatga
     2941 ccatgaaaat tcgtaacacc ggttagggcc cgaacaaaaa ctcatctcag aagaggatct
     3001 gaatagcgcc gtcgaccatc atcataatca tcattgagtt taaacccgct gatcagcctc
     3061 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     3121 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     3181 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     3241 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggaga tctatcgatg
     3301 ttctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag
     3361 ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     3421 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     3481 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     3541 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     3601 acgtacctag aagttcctat tccgaagttc ctattctcta gaaagtatag gaacttcctt
     3661 ggccaaaaag cctgaactca ccgcgacgtc tgtcgagaag tttctgatcg aaaagttcga
     3721 cagcgtctcc gacctgatgc agctctcgga gggcgaagaa tctcgtgctt tcagcttcga
     3781 tgtaggaggg cgtggatatg tcctgcgggt aaatagctgc gccgatggtt tctacaaaga
     3841 tcgttatgtt tatcggcact ttgcatcggc cgcgctcccg attccggaag tgcttgacat
     3901 tggggaattc agcgagagcc tgacctattg catctcccgc cgtgcacagg gtgtcacgtt
     3961 gcaagacctg cctgaaaccg aactgcccgc tgttctgcag ccggtcgcgg aggccatgga
     4021 tgcgatcgct gcggccgatc ttagccagac gagcgggttc ggcccattcg gaccgcaagg
     4081 aatcggtcaa tacactacat ggcgtgattt catatgcgcg attgctgatc cccatgtgta
     4141 tcactggcaa actgtgatgg acgacaccgt cagtgcgtcc gtcgcgcagg ctctcgatga
     4201 gctgatgctt tgggccgagg actgccccga agtccggcac ctcgtgcacg cggatttcgg
     4261 ctccaacaat gtcctgacgg acaatggccg cataacagcg gtcattgact ggagcgaggc
     4321 gatgttcggg gattcccaat acgaggtcgc caacatcttc ttctggaggc cgtggttggc
     4381 ttgtatggag cagcagacgc gctacttcga gcggaggcat ccggagcttg caggatcgcc
     4441 gcggctccgg gcgtatatgc tccgcattgg tcttgaccaa ctctatcaga gcttggttga
     4501 cggcaatttc gatgatgcag cttgggcgca gggtcgatgc gacgcaatcg tccgatccgg
     4561 agccgggact gtcgggcgta cacaaatcgc ccgcagaagc gcggccgtct ggaccgatgg
     4621 ctgtgtagaa gtactcgccg atagtggaaa ccgacgcccc agcactcgtc cgagggcaaa
     4681 ggaatagcac gtactacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     4741 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     4801 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     4861 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     4921 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     4981 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     5041 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     5101 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     5161 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     5221 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     5281 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     5341 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     5401 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     5461 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     5521 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     5581 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     5641 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     5701 gtccaacccg gtaagacacg acttatcgcc actggcagca gccac
//
