LOCUS       sequence-466387-        6861 bp    DNA     circular SYN 15-APR-2025
DEFINITION  PGK promoter driven FKBP fused with ADAR2dd (E488Q,T501A), with a 
            CD28 transmembrane domain in between.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6861)
  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 6861)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: YH039 PGK-TO-FKBP-tm-GS-XTEN-ADAR2(DD-E488Q-T5...
FEATURES             Location/Qualifiers
     source          1..6861
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      417..1005
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     906..925
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1159..1176
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1293..1314
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1329..1334,1335..1352,1353..1359)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1367..1383
                     /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     1372..1394
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1391..1407
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1391..1407
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    1456..1577
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1467..1486)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1521..1540
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1707..2729)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             complement(1707..2729)
                     /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
                     "
     protein_bind    complement(2737..2784)
                     /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)."
     primer_bind     2916..2935
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    3107..3331
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     3320..3337
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(3393..3422)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(join(4944..5264,5265..5267))
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="MGVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNK
                     PFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVEL
                     LKLE"
     regulatory      5333..5342
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(5377..5876)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(5425..5444)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     5818..5841
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     6048..6067
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        6142..6246
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6247..6315,6316..6861,1..246)
                     /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(6465..6484)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
       61 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
      121 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
      181 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
      241 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
      301 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
      361 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
      421 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
      481 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
      541 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
      601 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
      661 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
      721 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
      781 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
      841 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
      901 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
      961 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     1021 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     1081 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     1141 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     1201 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc
     1261 gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca
     1321 ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa
     1381 caatttcaca caggaaacag ctatgaccat gattacgcca agctctagct agaggtcgac
     1441 ggtatacaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
     1501 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
     1561 gctgcaataa acaagttggg gtgggcgaag aactccagca tgagatcccc gcgctggagg
     1621 atcatccagc cggcgtcccg gaaaacgatt ccgaagccca acctttcata gaaggcggcg
     1681 gtggaatcga aatctcgtag tacgtgctat tcctttgccc tcggacgagt gctggggcgt
     1741 cggtttccac tatcggcgag tacttctaca cagccatcgg tccagacggc cgcgcttctg
     1801 cgggcgattt gtgtacgccc gacagtcccg gctccggatc ggacgattgc gtcgcatcga
     1861 ccctgcgccc aagctgcatc atcgaaattg ccgtcaacca agctctgata gagttggtca
     1921 agaccaatgc ggagcatata cgcccggagc cgcggcgatc ctgcaagctc cggatgcctc
     1981 cgctcgaagt agcgcgtctg ctgctccata caagccaacc acggcctcca gaagaagatg
     2041 ttggcgacct cgtattggga atccccgaac atcgcctcgc tccagtcaat gaccgctgtt
     2101 atgcggccat tgtccgtcag gacattgttg gagccgaaat ccgcgtgcac gaggtgccgg
     2161 acttcggggc agtcctcggc ccaaagcatc agctcatcga gagcctgcgc gacggacgca
     2221 ctgacggtgt cgtccatcac agtttgccag tgatacacat ggggatcagc aatcgcgcat
     2281 atgaaatcac gccatgtagt gtattgaccg attccttgcg gtccgaatgg gccgaacccg
     2341 ctcgtctggc taagatcggc cgcagcgatc gcatccatgg cctccgcgac cggctgcaga
     2401 acagcgggca gttcggtttc aggcaggtct tgcaacgtga caccctgtgc acggcgggag
     2461 atgcaatagg tcaggctctc gctgaattcc ccaatgtcaa gcacttccgg aatcgggagc
     2521 gcggccgatg caaagtgccg ataaacataa cgatctttgt agaaaccatc ggcgcagcta
     2581 tttacccgca ggacatatcc acgccctcct acatcgaagc tgaaagcacg agattcttcg
     2641 ccctccgaga gctgcatcag gtcggagacg ctgtcgaact tttcgatcag aaacttctcg
     2701 acagacgtcg cggtgagttc aggctttttg gccaaggaag ttcctatact ttctagagaa
     2761 taggaacttc ggaataggaa cttctaggta cgtgaaccat caccctaatc aagttttttg
     2821 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     2881 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     2941 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     3001 aatgcgccgc tacagggcgc gtggggatac cccctagagc cccagctggt tctttccgcc
     3061 tcagaagcca tagagcccac cgcatcccca gaacatcgat agatctccat agagcccacc
     3121 gcatccccag catgcctgct attgtcttcc caatcctccc ccttgctgtc ctgccccacc
     3181 ccacccccca gaatagaatg acacctactc agacaatgcg atgcaatttc ctcattttat
     3241 taggaaagga cagtgggagt ggcaccttcc agggtcaagg aaggcacggg ggaggggcaa
     3301 acaacagatg gctggcaact agaaggcaca gtcgaggctg atcagcgggt ttaaactcaa
     3361 tgatgattat gatgatggtc gacggcgcta ttcagatcct cttctgagat gagtttttgt
     3421 tcgggccctc ctcgagcggc tcagggcgtg agtgagaact ggtcctgctc ggtgggcttc
     3481 tccacccagg cccccagccc cgccttgatg aaggctgtga acagacgcgc cttggcggcc
     3541 tggtactcct ttgccgccag cttggactca tggtacacgt tgggtttggt aatcttggag
     3601 cgtagtaagt gggagggaac cttgccgtgc acacgcatcc agcgacagta caacgcgtgc
     3661 ttacacaggc gggacgcgcg gcccagctca tccttcccag tcgtggcgtt gatgacctca
     3721 atagcggagt cgcctaccgt ccagttgaca ctgaagttgg gggccttccc tggctgccgt
     3781 gcttctgcat tgctgatgcc actgagcaaa ggcttgttga gggtgtagag aggtggcagg
     3841 tcctctatgt tggagatccg ctggtacatg gccctggaaa ggtggtcccc gtggtaaagg
     3901 ctgcccagga tgatgctcga gaagtaaatg ggctccacga aaatgctgag cagggatccc
     3961 tggatgccca ccacgttcca gcgtgcaatc ttgtcactgc aggacatggt gagcagccgc
     4021 tccccttgca gcaccccgtc ccacgcttgg atgctcgcat tggagcgcac tggaatcgtc
     4081 ccctgaccag actctatttt ggtccgtagc tgtcctcttg ctttacgatt tgggtgtcta
     4141 tctgctggtt cttccaggat tggctcatgt ggtgagaaga ttctggcatc tccacaggga
     4201 gaggtgctga tgtacagatg aaactggaca ttctccttca gcctaaaccc ccctcgctct
     4261 gatttctgaa agatggatct tttttgatca tctttgttat ttaagtaaag ctcaagttgt
     4321 gtataaagaa atctgagcaa ggatctccga gatattattt ctgcatggca gtcatttaat
     4381 gcaaggccac gatcactcat gtattcacca ttaatacatt ttgttcctgt agaaacactt
     4441 atcaccttgg catctttaac atctgtgcct gttgtcatga cgactccagc cagcactttt
     4501 ctgcgagcgt gaggggagga gaagttgtcg gtcaggtcac caaacttacc caggaccagg
     4561 cgtgagacag cgtcagctaa aacctgcggt aaatgcagct gaagaccctc actgggaata
     4621 ggctggcgag atggcgtttg atccaagtgc aaagcgctag cactctctgg cgttgctgac
     4681 tcactcgtgc cgggtgtttc tgacccactg gacccgctac ccgccccgct gccggaaccg
     4741 cctgacccgg aattttcaac acaaagagcc actgtcacta gcaagccata acaaaacagg
     4801 actccagcaa ccacgaccag accggtacct ccccccgaac cgccactacc accagacccc
     4861 cccgatccgc cgttgtcgtc atcgtccttg tagtctcctc cagatccgcc agtgccaccc
     4921 ccagaccctc ctgtcccggc gccttccagt tttagaagct ccacatcgaa gacgagagtg
     4981 gcatgtggtg ggatgatgcc tgggtgccca gtggcaccat aggcataatc tggagatata
     5041 gtcagtttgg ctctctgacc cacactcatc tgggcaaccc cttcttccca gcctcggatc
     5101 acctcctgct tgcctagcat aaacttaaag ggcttgtttc tgtcccggga ggaatcaaat
     5161 ttctttccat cttcaagcat cccggtgtag tgcaccacgc aggtctggcc gcgcttgggg
     5221 aaggtgcgcc cgtctcctgg ggagatggtt tccacctgca ctcccatgga tccaggccgt
     5281 gcagcgtgca gcagcagtgc cagcggaaga agcagtgcgg tgacagggag cgccatggtg
     5341 gcgaagctta agtttaaacg ctagagtcct gcaggtcgaa aggcccggag atgaggaaga
     5401 ggagaacagc gcggcagacg tgcgcttttg aagcgtgcag aatgccgggc ctccggagga
     5461 ccttcgggcg cccgccccgc ccctgagccc gcccctgagc ccgcccccgg acccacccct
     5521 tcccagcctc tgagcccaga aagcgaagga gcaaagctgc tattggccgc tgccccaaag
     5581 gcctacccgc ttccattgct cagcggtgct gtccatctgc acgagactaa tgagacgtgc
     5641 tacttccatt tgtcacgtcc tgcacgacgc gagctgcggg gcggggggga acttcctgac
     5701 taggggagga gtagaaggtg gcgcgaaggg gccaccaaag aacggagccg gttggcgcct
     5761 accggtggat gtggaatgtg tgcgaggcca gaggccactt gtgtagcgcc aagtgcccag
     5821 cggggctgct aaagcgcatg ctccagactg ccttgggaaa agcgcctccc ctacccggta
     5881 gaacgcgtat atctggcccg tacgaagcag cgcaaaacgc ctaaccctaa gcagattctt
     5941 catgcaattg tcggtcaagc cttgccttgt tgtagcttaa attttgctcg cgcactactc
     6001 agcgacctcc aacacacaag cagggagcag atactggctt aactatgcgg catcagagca
     6061 gattgtactg agagtgcacc ataggggatc gggagatctc ccgatccgtc gacgtcaggt
     6121 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     6181 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     6241 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     6301 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     6361 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     6421 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     6481 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     6541 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     6601 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     6661 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     6721 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     6781 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     6841 ctagcttccc ggcaacaatt a
//
