LOCUS       sequence-417083-        6825 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient mammalian expression of the CC functionalized GEMS 
            receptor A-a3-GEMS (JAK-STAT).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6825)
  AUTHORS   Makri Pistikou AM, Cremers GAO, Nathalia BL, Meuleman TJ, Bogels 
            BWA, Eijkens BV, de Dreu A, Bezembinder MTH, Stassen OMJA, Bouten 
            CCV, Merkx M, Jerala R, de Greef TFA
  TITLE     Engineering a scalable and orthogonal platform for synthetic 
            communication in mammalian cells.
  JOURNAL   Nat Commun. 2023 Nov 2;14(1):7001. doi: 10.1038/s41467-023-42810-5.
  PUBMED    37919273
REFERENCE   2  (bases 1 to 6825)
  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 
            Commun."; date: "2023-11-2"; volume: "14(1):7001. doi"; pages: " 
            10.1038/s41467-023-42810-5"
COMMENT     Sequence Label: pLeo-A-a3-GEMS(JAK-STAT)
FEATURES             Location/Qualifiers
     source          1..6825
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(49..68)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     659..678
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    963..1084
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1000..1019)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1054..1073
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1133..1149)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1133..1149)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1146..1168)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1157..1173
                     /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(1181..1187,1188..1205,1206..1211))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1226..1247
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1364..1381)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1535..2123)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1615..1634)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2294..3085,3086..3154))
                     /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     2917..2936
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3155..3259)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3334..3353)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        3535..3731
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3582..3717
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      3757..3766
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3832..3858
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(5706..5723)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5712..5936
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5982..6410
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6069..6088)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6279..6300
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6419..6439)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6424..6753
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6604..6739
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             join(6820..6825,1..789)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
ORIGIN
        1 gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat
       61 gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag
      121 gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac
      181 gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac
      241 gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc
      301 ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg
      361 ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag
      421 cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat
      481 caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag
      541 gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc
      601 ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg
      661 ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg
      721 ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag
      781 ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat
      841 cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc
      901 gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc
      961 ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     1021 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     1081 cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc
     1141 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     1201 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     1261 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     1321 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     1381 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     1441 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     1501 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     1561 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     1621 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     1681 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     1741 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     1801 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     1861 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     1921 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     1981 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     2041 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     2101 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     2161 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     2221 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     2281 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     2341 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     2401 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     2461 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     2521 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     2581 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     2641 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     2701 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     2761 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     2821 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     2881 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     2941 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     3001 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     3061 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     3121 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     3181 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     3241 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg
     3301 gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     3361 aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat
     3421 ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc ttagggttag
     3481 gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgtgatctgc gatctgcatc
     3541 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     3601 ccagttccgc ccattctccg ccccatcgct gactaatttt ttttatttat gcagaggccg
     3661 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     3721 gcttttgcaa aaagcttcga atcgcgctag cggccggcca ccatggaaac tgatactttg
     3781 ctgctctggg tcctgctgct gtgggtccct ggatcaacgg gggacggatc ctacccatac
     3841 gatgttccag attacgctgg tacctccggc cctggcagca gccccgagga cgagatccag
     3901 caactggaag aggaaatcgc ccagctggag cagaaaaacg ccgctctgaa ggaaaagaac
     3961 caggccctga agtacgaatt cattcgtagt ggatcctcgg gatcatcagg atcctctggg
     4021 agctcaggat cctcggggag ctctggatcc agcggttcaa gcggatccac tagtgcacct
     4081 tcacccagcc tcccggaccc caagtttgag agcaaagcgg ccctgctggc atcccggggc
     4141 tccgaagaac ttctgtgctt cacccaacgc ttggaagact tggtgtgttt ctgggaggaa
     4201 gcggcgagct ccgggatgga cttcaactac agcttctcat accagctcga gggtgagtca
     4261 cgaaagtcat gtagcctgca ccaggctccc accgtccgcg gctccgtgcg tttctggtgt
     4321 tcactgccaa cagcggacac atcgagtgct gtgccgctgg agctgcaggt gacggaggcg
     4381 tccggttctc ctcgctatca ccgcatcatc catatcaatg aagtagtgct cctggacgcc
     4441 cccgcggggc tgctggcgcg ccgggcagaa gagggcagcc acgtggtgct gcgctggctg
     4501 ccacctcctg gagcacctat gaccacccac atccgatatg aagtggacgt gtcggcaggc
     4561 aaccgggcag gagggacaca aagggtggag gtcctggaag gccgcactga gtgtgttctg
     4621 agcaacctgc ggggcgggac gcgctacacc ttcgctgttc gagcgcgcat ggccgagccg
     4681 agcttcagcg gattctggag tgcctggtct gagcccgcgt cactactgac cgctagcgac
     4741 ctggaccctc tcatcttgac gctgtctctc attctggtcc tcatctcgtt gttgctgacg
     4801 gttctggccc tgctgtccgc agcagcaaat aagcgagacc taattaaaaa acacatctgg
     4861 cctaatgttc cagatccttc aaagagtcat attgcccagt ggtcacctca cactcctcca
     4921 aggcacaatt ttaattcaaa agatcaaatg tattcagatg gcaatttcac tgatgtaagt
     4981 gttgtggaaa tagaagcaaa tgacaaaaag ccttttccag aagatctgaa atcattggac
     5041 ctgttcaaaa aggaaaaaat taatactgaa ggacacagca gtggtattgg ggggtcttca
     5101 tgcatgtcat cttctaggcc aagcatttct agcagtgatg aaaatgaatc ttcacaaaac
     5161 acttcgagca ctgtccaggc ttctaccgtg gtacacagtg gctacagaca ccaagttccg
     5221 tcagtccaag tcttctcaag atccgagtct acccagccct tgttagattc agaggagcgg
     5281 ccagaagatc tacaattagt agatcatgta gatggcggtg atggtatttt gcccaggcaa
     5341 cagtacttca aacagaactg cagtcagcat gaatccagtc cagatatttc acattttgaa
     5401 aggtcaaagc aagtttcatc agtcaatgag gaagattttg ttagacttaa acagcagatt
     5461 tcagatcata tttcacaatc ctgtggatct gggcaaatga aaatgtttca ggaagtttct
     5521 gcagcagatg cttttggtcc aggtactgag ggacaagtag aaagatttga aacagttggc
     5581 atggaggctg cgactgatga aggcatgcct aaaagttact taccacagac tgtacggcaa
     5641 ggcggctaca tgcctcagta ggcggccgct cgagtctaga gggcccgttt aaacccgctg
     5701 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     5761 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     5821 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     5881 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     5941 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     6001 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     6061 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     6121 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     6181 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     6241 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     6301 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     6361 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     6421 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     6481 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     6541 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     6601 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     6661 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     6721 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     6781 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgatt
//
