LOCUS       sequence-467416-        6496 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  plasmid expressing membrane-bound CCL20 by fusing with pre-mGRASP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6496)
  AUTHORS   Zhang X, Mille-Fragoso LS, Kaseniit KE, Lee AP, Zhang M, Call CC, Hu
            Y, Xie Y, Gao XJ
  TITLE     Post-transcriptional modular synthetic receptors.
  JOURNAL   Nat Chem Biol. 2025 Mar 28. doi: 10.1038/s41589-025-01872-w.
  PUBMED    40155716
REFERENCE   2  (bases 1 to 6496)
  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 Mar 28. doi: 10.1038/s41589-025-01872-w."
COMMENT     Sequence Label: XZ033 CMV-TO-CCL20-pre-mGRASP
FEATURES             Location/Qualifiers
     source          1..6496
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        384..587
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     538..558
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    589..607
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    610..628
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     638..662
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1135..1224
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     CDS             2098..2127
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(2183..2200)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2189..2413
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2585..2604)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2736..2783
                     /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             2791..3813
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2791..3813
                     /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    3943..4064
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3980..3999)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4034..4053
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4113..4129)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4113..4129)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4126..4148)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4137..4153
                     /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(4161..4167,4168..4185,4186..4191))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4206..4227
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4344..4361)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4515..5103)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4595..4614)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5274..6065,6066..6134))
                     /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     5897..5916
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6135..6239)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6314..6333)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gttgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
      421 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
      481 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
      541 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctctc cctatcagtg
      601 atagagatct ccctatcagt gatagagatc gtcgacgagc tcgtttagtg aaccgtcaga
      661 tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccgg gaccgatcca
      721 gcctccggac tctagcgttt aaacttaagc ttcgccacca tgtgctgtac caagagtttg
      781 ctcctggctg ctttgatgtc agtgctgcta ctccacctct gcggcgaatc agaagcagca
      841 agcaactttg actgctgtct tggatacaca gaccgtattc ttcatcctaa atttattgtg
      901 ggcttcacac ggcagctggc caatgaaggc tgtgacatca atgctatcat ctttcacaca
      961 aagaaaaagt tgtctgtgtg cgcaaatcca aaacagactt gggtgaaata tattgtgcgt
     1021 ctcctcagta aaaaagtcaa gaacatgggt accgggaacg gaaatggcaa tggtaacggg
     1081 aacggaaatg gcaatggtaa cgggaatact agtgttaaca ccggtgcggc cgtttaccca
     1141 tacgatgttc ctgactatgc gggctatccc tatgacgtcc cggactatgc aggatcctat
     1201 ccatatgacg ttccagatta cgctccggcc gccaccggtc ttaaggtcga ctttcagaaa
     1261 gctagctcta ttgtttataa gaaggagggg gaacaggtgg agttcagttt cccattggcc
     1321 ttcacagtcg agaagctcac aggatcagga gagttgtggt ggcaggccga aagagcctcc
     1381 tccagcaaga gttggatcac ttttgatctg aaaaacaaag aagtgagcgt gaagagagtc
     1441 acacaggacc ctaagctgca gatgggcaag aagttgcctt tgcacttgac actcccccaa
     1501 gccctgcccc agtatgctgg ctccgggaac ctcaccctcg ccttggaggc taagaccgga
     1561 aagttgcatc aggaggtcaa cctcgtggtg atgagggcaa cacagctgca gaagaacctg
     1621 acctgcgaag tgtggggtcc cacctcacct aaactgatgc tgtctctgaa gctggagaat
     1681 aaggaggcta aagtcagtaa aagagagaag gctgtgtggg tccttaaccc ggaggcaggg
     1741 atgtggcagt gtcttcttag cgactctgga caggtgctgc tggagagcaa catcaaggtg
     1801 cttcctactt ggtcaacacc agtacagccg atggccctga tcgtactcgg gggagtcgcc
     1861 ggcctgcttc tgtttatcgg cctgggcatc ttcttctgcg tacgctacaa gtatcgcaac
     1921 agggatgagg gatcttacca cgttgacgaa tctaggaact atatcagtaa cagcgctcag
     1981 agtaatggcg ccgtggttaa agagaaacag ccctccagcg ccaaatcagc aaacaaaaac
     2041 aagaaaaaca aggataaaga gtactacgta cttaagtaag ccgctcgagg agggcccgaa
     2101 caaaaactca tctcagaaga ggatctgaat agcgccgtcg accatcatca taatcatcat
     2161 tgagtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     2221 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     2281 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     2341 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     2401 gtgggctcta tggagatcta tcgatgttct ggggatgcgg tgggctctat ggcttctgag
     2461 gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta
     2521 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     2581 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     2641 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     2701 aaaaaacttg attagggtga tggttcacgt acctagaagt tcctattccg aagttcctat
     2761 tctctagaaa gtataggaac ttccttggcc aaaaagcctg aactcaccgc gacgtctgtc
     2821 gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct ctcggagggc
     2881 gaagaatctc gtgctttcag cttcgatgta ggagggcgtg gatatgtcct gcgggtaaat
     2941 agctgcgccg atggtttcta caaagatcgt tatgtttatc ggcactttgc atcggccgcg
     3001 ctcccgattc cggaagtgct tgacattggg gaattcagcg agagcctgac ctattgcatc
     3061 tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg aaaccgaact gcccgctgtt
     3121 ctgcagccgg tcgcggaggc catggatgcg atcgctgcgg ccgatcttag ccagacgagc
     3181 gggttcggcc cattcggacc gcaaggaatc ggtcaataca ctacatggcg tgatttcata
     3241 tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg tgatggacga caccgtcagt
     3301 gcgtccgtcg cgcaggctct cgatgagctg atgctttggg ccgaggactg ccccgaagtc
     3361 cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa tggccgcata
     3421 acagcggtca ttgactggag cgaggcgatg ttcggggatt cccaatacga ggtcgccaac
     3481 atcttcttct ggaggccgtg gttggcttgt atggagcagc agacgcgcta cttcgagcgg
     3541 aggcatccgg agcttgcagg atcgccgcgg ctccgggcgt atatgctccg cattggtctt
     3601 gaccaactct atcagagctt ggttgacggc aatttcgatg atgcagcttg ggcgcagggt
     3661 cgatgcgacg caatcgtccg atccggagcc gggactgtcg ggcgtacaca aatcgcccgc
     3721 agaagcgcgg ccgtctggac cgatggctgt gtagaagtac tcgccgatag tggaaaccga
     3781 cgccccagca ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt cgattccacc
     3841 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     3901 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
     3961 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     4021 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     4081 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     4141 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     4201 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     4261 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     4321 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     4381 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     4441 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     4501 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4561 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4621 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4681 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     4741 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     4801 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     4861 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     4921 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     4981 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     5041 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     5101 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     5161 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     5221 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     5281 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     5341 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     5401 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     5461 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     5521 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     5581 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     5641 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     5701 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     5761 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     5821 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     5881 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     5941 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     6001 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     6061 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     6121 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     6181 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     6241 acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat
     6301 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg
     6361 cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag
     6421 gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgta
     6481 cgggccagat atacgc
//
