LOCUS       sequence-466408-        6246 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PGK promoter driven FRB fused with MCP, with a CD28 transmembrane 
            domain in between.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6246)
  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 6246)
  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: YH038 PGK-TO-FRB-tm-MCP
FEATURES             Location/Qualifiers
     source          1..6246
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      265..274
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(309..808)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(357..376)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     750..773
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     980..999
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        1074..1178
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1179..1247,1248..2039)
                     /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(1397..1416)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2210..2798
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2699..2718
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2952..2969
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3086..3107
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3122..3127,3128..3145,3146..3152)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3160..3176
                     /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     3165..3187
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3184..3200
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3184..3200
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    3249..3370
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3260..3279)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3314..3333
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(3500..4522)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             complement(3500..4522)
                     /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(4530..4577)
                     /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     4709..4728
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    4900..5124
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     5113..5130
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(5186..5215)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
ORIGIN
        1 tcatgtactt cctgcaccac tcttgggcct ccattaaatc tcgaccatag gcctgattaa
       61 aggatgtttc cttcagagtc tggggtcccc gttccatcat agcatgcaag ggctccagca
      121 cctcaaacat gcctttcacg ttcctttccc caaagtacaa acgagatgcc tcttccaggc
      181 cttcatgcca catctccagg gatccaggcc gtgcagcgtg cagcagcagt gccagcggaa
      241 gaagcagtgc ggtgacaggg agcgccatgg tggcgaagct taagtttaaa cgctagagtc
      301 ctgcaggtcg aaaggcccgg agatgaggaa gaggagaaca gcgcggcaga cgtgcgcttt
      361 tgaagcgtgc agaatgccgg gcctccggag gaccttcggg cgcccgcccc gcccctgagc
      421 ccgcccctga gcccgccccc ggacccaccc cttcccagcc tctgagccca gaaagcgaag
      481 gagcaaagct gctattggcc gctgccccaa aggcctaccc gcttccattg ctcagcggtg
      541 ctgtccatct gcacgagact aatgagacgt gctacttcca tttgtcacgt cctgcacgac
      601 gcgagctgcg gggcgggggg gaacttcctg actaggggag gagtagaagg tggcgcgaag
      661 gggccaccaa agaacggagc cggttggcgc ctaccggtgg atgtggaatg tgtgcgaggc
      721 cagaggccac ttgtgtagcg ccaagtgccc agcggggctg ctaaagcgca tgctccagac
      781 tgccttggga aaagcgcctc ccctacccgg tagaacgcgt atatctggcc cgtacgaagc
      841 agcgcaaaac gcctaaccct aagcagattc ttcatgcaat tgtcggtcaa gccttgcctt
      901 gttgtagctt aaattttgct cgcgcactac tcagcgacct ccaacacaca agcagggagc
      961 agatactggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatagggga
     1021 tcgggagatc tcccgatccg tcgacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     1081 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     1141 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     1201 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     1261 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     1321 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     1381 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     1441 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     1501 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     1561 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     1621 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     1681 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     1741 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     1801 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     1861 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     1921 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     1981 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     2041 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     2101 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     2161 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     2221 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     2281 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     2341 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     2401 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     2461 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     2521 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     2581 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     2641 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     2701 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     2761 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     2821 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     2881 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     2941 cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc
     3001 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     3061 aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg
     3121 ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc
     3181 acacaggaaa cagctatgac catgattacg ccaagctcta gctagaggtc gacggtatac
     3241 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     3301 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     3361 taaacaagtt ggggtgggcg aagaactcca gcatgagatc cccgcgctgg aggatcatcc
     3421 agccggcgtc ccggaaaacg attccgaagc ccaacctttc atagaaggcg gcggtggaat
     3481 cgaaatctcg tagtacgtgc tattcctttg ccctcggacg agtgctgggg cgtcggtttc
     3541 cactatcggc gagtacttct acacagccat cggtccagac ggccgcgctt ctgcgggcga
     3601 tttgtgtacg cccgacagtc ccggctccgg atcggacgat tgcgtcgcat cgaccctgcg
     3661 cccaagctgc atcatcgaaa ttgccgtcaa ccaagctctg atagagttgg tcaagaccaa
     3721 tgcggagcat atacgcccgg agccgcggcg atcctgcaag ctccggatgc ctccgctcga
     3781 agtagcgcgt ctgctgctcc atacaagcca accacggcct ccagaagaag atgttggcga
     3841 cctcgtattg ggaatccccg aacatcgcct cgctccagtc aatgaccgct gttatgcggc
     3901 cattgtccgt caggacattg ttggagccga aatccgcgtg cacgaggtgc cggacttcgg
     3961 ggcagtcctc ggcccaaagc atcagctcat cgagagcctg cgcgacggac gcactgacgg
     4021 tgtcgtccat cacagtttgc cagtgataca catggggatc agcaatcgcg catatgaaat
     4081 cacgccatgt agtgtattga ccgattcctt gcggtccgaa tgggccgaac ccgctcgtct
     4141 ggctaagatc ggccgcagcg atcgcatcca tggcctccgc gaccggctgc agaacagcgg
     4201 gcagttcggt ttcaggcagg tcttgcaacg tgacaccctg tgcacggcgg gagatgcaat
     4261 aggtcaggct ctcgctgaat tccccaatgt caagcacttc cggaatcggg agcgcggccg
     4321 atgcaaagtg ccgataaaca taacgatctt tgtagaaacc atcggcgcag ctatttaccc
     4381 gcaggacata tccacgccct cctacatcga agctgaaagc acgagattct tcgccctccg
     4441 agagctgcat caggtcggag acgctgtcga acttttcgat cagaaacttc tcgacagacg
     4501 tcgcggtgag ttcaggcttt ttggccaagg aagttcctat actttctaga gaataggaac
     4561 ttcggaatag gaacttctag gtacgtgaac catcacccta atcaagtttt ttggggtcga
     4621 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     4681 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     4741 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     4801 cgctacaggg cgcgtgggga taccccctag agccccagct ggttctttcc gcctcagaag
     4861 ccatagagcc caccgcatcc ccagaacatc gatagatctc catagagccc accgcatccc
     4921 cagcatgcct gctattgtct tcccaatcct cccccttgct gtcctgcccc accccacccc
     4981 ccagaataga atgacaccta ctcagacaat gcgatgcaat ttcctcattt tattaggaaa
     5041 ggacagtggg agtggcacct tccagggtca aggaaggcac gggggagggg caaacaacag
     5101 atggctggca actagaaggc acagtcgagg ctgatcagcg ggtttaaact caatgatgat
     5161 tatgatgatg gtcgacggcg ctattcagat cctcttctga gatgagtttt tgttcgggcc
     5221 ctcctcgagc ggctcagtaa attccgctat tcgcagcgat tgcggaaggt atcgggttgc
     5281 catctttcag aagtccttgc atggctttta caataagctc gcagtcgcta ttggtcgcaa
     5341 atatcggaat cgtcaattcc atattcaagt atgagcgcca tgcacctttg gggacctcca
     5401 ctttaatggt atacttccgg ttctgggcac tagattggcg tacactacaa gtcaccttat
     5461 aggcttgtga tcttgaattg gaagagatcc actcggctat cccattcgca aagttacttg
     5521 gagctactgt gacgtctcca gtgccgccgt tgtccacgag cacaaactgt gtaaaattgg
     5581 aggccatgga cccgccaccg gttccgccac catatattcc tgagttggcg gcgatcgcag
     5641 agggaatcgg gttaccgtct ttcaaaagac cttgcatggc cttgacaatc agctcgcagt
     5701 ctgaattagt cgcaaagatt gggatcgtaa gctccatatt aagataagat cgccacgccc
     5761 cttttggcac ttctaccttt atggtatatt ttcggttctg tgcggaactt tggcgaactg
     5821 agcatgtgac tttatatgcc tgactccggc tgttactact tatccattct gcaatcccgt
     5881 tcgcaaaatt gctcggcgca acagttacgt ctccggtgcc accattgtca accaacacaa
     5941 attgcgtaaa gttgctcgcc atagcgctac caccgccgtt ttcaacacaa agagccactg
     6001 tcactagcaa gccataacaa aacaggactc cagcaaccac gaccagaccg gtacctcccc
     6061 ccgaaccgcc actaccacca gacccccccg atccgccgtt gtcgtcatcg tccttgtagt
     6121 ctcctccaga tccgccagtg ccacccccag accctcctgt cccggcgcct ctagtctttg
     6181 agattcgtcg gaacacatga taatagaggt cccaggcttg gaggaggtcc ttgacattcc
     6241 ctgatt
//
