LOCUS       sequence-266807-        6607 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses both the gePSI-alpha-chain and the gePSI-beta-chain under 
            control of the inducible bi-directional TRE3G promoter. Expresses 
            constitutively AcGFP under control of a CMV promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6607)
  AUTHORS   Heumuller M, Glock C, Rangaraju V, Biever A, Schuman EM
  TITLE     A genetically encodable cell-type-specific protein synthesis 
            inhibitor.
  JOURNAL   Nat Methods. 2019 Jul 15. pii: 10.1038/s41592-019-0468-x. doi: 
            10.1038/s41592-019-0468-x.
  PUBMED    31308551
REFERENCE   2  (bases 1 to 6607)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Methods. 2019 Jul 15. pii: 10.1038/s41592-019-0468-x. doi: 
            10.1038/s41592-019-0468-x."
COMMENT     Sequence Label: pTRE3G-Bi-gePSI-pCMV-AcGFP
FEATURES             Location/Qualifiers
     source          1..6607
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..451
                     /label=TRE3G BI promoter
                     /note="3rd-generation Tet-responsive bidirectional promoter
                     that can be activated by binding of Tet-On(R) 3G"
     protein_bind    81..99
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    117..135
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    153..171
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    189..207
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    225..243
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    261..279
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    297..315
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     polyA_signal    1031..1112
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     enhancer        1223..1602
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1603..1806
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1757..1777
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1851..1870
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1851..1869
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1915..2634
                     /codon_start=1
                     /product="Aequorea coerulescens GFP"
                     /label=AcGFP1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFEDD
                     GNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMIYF
                     GFVTAAAITHGMDELYK"
     primer_bind     complement(2649..2666)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2655..2879
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2964..3104
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2964..3093
                     /label=AAV2 ITR
     rep_origin      3179..3634
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3266..3285)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3476..3497
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3651..3670)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3770..3792
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3830..3848)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3916..4020
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4021..4089,4090..4881)
                     /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(4239..4258)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5052..5640
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5541..5560
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   5702..5831
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     polyA_signal    5967..6048
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     regulatory      6586..6595
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 gcgatctgac ggttcactaa acgagctctg cttatatagg cctcccaccg tacacgccac
       61 ctcgacatac tcgagtttac tccctatcag tgatagagaa cgtatgaaga gtttactccc
      121 tatcagtgat agagaacgta tgcagacttt actccctatc agtgatagag aacgtataag
      181 gagtttactc cctatcagtg atagagaacg tatgaccagt ttactcccta tcagtgatag
      241 agaacgtatc tacagtttac tccctatcag tgatagagaa cgtatatcca gtttactccc
      301 tatcagtgat agagaacgta taagctttag gcgtgtacgg tgggcgccta taaaagcaga
      361 gctcgtttag tgaaccgtca gatcgcctgg agcaattcca caacactttt gtcttatacc
      421 aactttccgt accacttcct accctcgtaa agtcgacacc ggggcccaga tctatgaagt
      481 tcactgaaat cttccccgtg gaggacgcga actaccctta cagcgccttc atcgcgtcgg
      541 tccggaaaga cgtgatcaaa cactgcaccg accataaagg gatcttccag cccgtgctgc
      601 caccggagaa gaaggtcccg gagctatggt tgtacacaga gctcaaaact aggaccagct
      661 ccatcacgct cgccatacgc atggacaacc tgtacctcgt gggcttcagg accccgggcg
      721 gggtgtggtg ggagttcggc aaggacggcg acacccacct cctcggcgac aaccccaggt
      781 ggctcggctt cggcggcagg taccaggacc tcatcggcaa caagggtctg gagaccgtca
      841 ccatgggccg cgccgaaatg accagggccg tcaacgacct ggcgaagaag aagaagtgag
      901 cggccgccgg cgatatctcc agaggatcat aatcagccat accacatttg tagaggtttt
      961 acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat
     1021 tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     1081 aaatttcaca aataaagcat ttttttcact gccccgagct tcctcgctca ctgactcgct
     1141 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaagggct gcttcgcgat
     1201 gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt
     1261 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     1321 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     1381 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     1441 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     1501 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     1561 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     1621 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     1681 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     1741 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     1801 agagctctct ggctaactag agaacccact gcttactggc ttatcgaaat taatacgact
     1861 cactataggg agacccaagc tggctagcgt ttaaacttaa gcttggtacc gagcatggtg
     1921 agcaagggcg ccgagctgtt caccggcatc gtgcccatcc tgatcgagct gaatggcgat
     1981 gtgaatggcc acaagttcag cgtgagcggc gagggcgagg gcgatgccac ctacggcaag
     2041 ctgaccctga agttcatctg caccaccggc aagctgcctg tgccctggcc caccctggtg
     2101 accaccctga gctacggcgt gcagtgcttc tcacgctacc ccgatcacat gaagcagcac
     2161 gacttcttca agagcgccat gcctgagggc tacatccagg agcgcaccat cttcttcgag
     2221 gatgacggca actacaagtc gcgcgccgag gtgaagttcg agggcgatac cctggtgaat
     2281 cgcatcgagc tgaccggcac cgatttcaag gaggatggca acatcctggg caataagatg
     2341 gagtacaact acaacgccca caatgtgtac atcatgaccg acaaggccaa gaatggcatc
     2401 aaggtgaact tcaagatccg ccacaacatc gaggatggca gcgtgcagct ggccgaccac
     2461 taccagcaga atacccccat cggcgatggc cctgtgctgc tgcccgataa ccactacctg
     2521 tccacccaga gcgccctgtc caaggacccc aacgagaagc gcgatcacat gatctacttc
     2581 ggcttcgtga ccgccgccgc catcacccac ggcatggatg agctgtacaa gtgaaacccg
     2641 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     2701 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     2761 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     2821 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     2881 ttctgaggcg gaaagaacca gatcctctct taaggtagca tcgagattta aattagggat
     2941 aacagggtaa tggcgcgggc cgcaggaacc cctagtgatg gagttggcca ctccctctct
     3001 gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc
     3061 ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc tgatgcggta
     3121 ttttctcctt acgcatctgt gcggtatttc acaccgcata cgtcaaagca accatagtac
     3181 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     3241 acacttgcca gcgccttagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     3301 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     3361 gctttacggc acctcgaccc caaaaaactt gatttgggtg atggttcacg tagtgggcca
     3421 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     3481 ctcttgttcc aaactggaac aacactcaac tctatctcgg gctattcttt tgatttataa
     3541 gggattttgc cgatttcggt ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     3601 gcgaatttta acaaaatatt aacgtttaca attttatggt gcactctcag tacaatctgc
     3661 tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga
     3721 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     3781 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata
     3841 cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact
     3901 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     3961 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     4021 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     4081 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     4141 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     4201 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     4261 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     4321 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     4381 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     4441 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     4501 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     4561 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     4621 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     4681 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     4741 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     4801 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     4861 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     4921 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     4981 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     5041 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     5101 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     5161 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     5221 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     5281 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     5341 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     5401 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     5461 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     5521 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     5581 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     5641 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     5701 tcctgcaggc agctgcgcgc tcgctcgctc actgaggccg cccgggcgtc gggcgacctt
     5761 tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca actccatcac
     5821 taggggttcc tgcggcaatt cagtcgataa ctataacggt cctaaggtag cgatttaaat
     5881 acgcgctctc ttaaggtagc cccgggacgc gtcaattgca tgaagaatct gcttagggtt
     5941 aggcgttttg ctgccacctg acgtcggcag tgaaaaaaat gctttatttg tgaaatttgt
     6001 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat
     6061 tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa
     6121 aacctctaca aatgtggtat ggctgattat gatcctctag actgcagact agttcattga
     6181 aagtacaagt tctctgaacc agcttttttg tagagagatg ttatgctttc ctgggcacaa
     6241 gacattggca gcttgtaaag ttcgtccatt ccgtgtgtta tcccagccgc cgcagtagtt
     6301 tgattcttaa cgagcgcaac gatcctcgct gcttcgttct tatccttgat gccaagcttc
     6361 tgcatgtcgg ggatcacagc ggtggggtgg tcagcccact cgaaggccgc cttggagatc
     6421 ctgtcccact tctgcacctg cttcccctgc gtcacggtca aggtcacccc gtgctggctg
     6481 ttgaaccccg cgtccaccgt gcgggacacg gtgttgaacc gcagcccctc gcacaccatg
     6541 accaccagct tcaccagctt gctcttcgtg tcggcctgtg ggtcagccgc catggtgaat
     6601 tctccag
//
