LOCUS       sequence-357259-        7935 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses TetOn3G transcription factor selectable by hygromycin 
            resistance (Lentiviral vector).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7935)
  AUTHORS   Gemperle J, Harrison TS, Flett C, Adamson AD, Caswell PT
  TITLE     On demand expression control of endogenous genes with DExCon, 
            DExogron and LUXon reveals differential dynamics of Rab11 family 
            members.
  JOURNAL   Elife. 2022 Jun 16;11. pii: 76651. doi: 10.7554/eLife.76651.
  PUBMED    35708998
REFERENCE   2  (bases 1 to 7935)
  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: "Elife."; 
            date: "2022-06-16"; volume: "11. pii: 76651. doi"; pages: " 
            10.7554/eLife.76651"
COMMENT     Sequence Label: pCDH-EF1a-mTagBFP-T2A-TetOn3G
FEATURES             Location/Qualifiers
     source          1..7935
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        6..232
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             233..413
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    460..585
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1078..1311
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1496..1540
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    1807..1923
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1968..2179
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             2192..2460
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     CDS             2493..3191
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     CDS             3201..3254
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3261..4007
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     misc_feature    4014..4602
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4067..4087)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4485..4496)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4676..4909
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4981..5102
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5018..5037)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5072..5091
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5142..5277
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5310..5326)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5310..5326)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5323..5345)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5334..5350
                     /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(5358..5364,5365..5382,5383..5388))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5403..5424
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5541..5558)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5712..6300)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6471..7262,7263..7331))
                     /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     7094..7113
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7332..7436)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7504..7522
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7560..7582)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7682..7701
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7895..7917
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7909..7926
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7910..7926
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 acgcgtgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga tgagttagca
       61 acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag taaggtggta
      121 cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac gaaccactga
      181 attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacataa acgggtctct
      241 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
      301 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
      361 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc
      421 ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg caggactcgg
      481 cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac gccaaaaatt
      541 ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat taagcggggg
      601 agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa
      661 ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa tcctggcctg
      721 ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc ccttcagaca
      781 ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg tgtgcatcaa
      841 aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga gcaaaacaaa
      901 agtaagacca ccgcacagca agcggccact gatcttcaga cctggaggag gagatatgag
      961 ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac cattaggagt
     1021 agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag tgggaatagg
     1081 agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag cgtcaatgac
     1141 gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga acaatttgct
     1201 gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca tcaagcagct
     1261 ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc tggggatttg
     1321 gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta gttggagtaa
     1381 taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca gagaaattaa
     1441 caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc aagaaaagaa
     1501 tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt ttaacataac
     1561 aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg taggtttaag
     1621 aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt caccattatc
     1681 gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa tagaagaaga
     1741 aggtggagag agagacagag acagatccat tcgattagtg aacggatctc gacggtatcg
     1801 gttaactttt aaaagaaaag gggggattgg ggggtacagt gcaggggaaa gaatagtaga
     1861 cataatagca acagacatac aaactaaaga attacaaaaa caaattacaa aattcaaaat
     1921 tttatcgaat tctaaaagga tctgcgatcg ctccggtgcc cgtcagtggg cagagcgcac
     1981 atcgcccaca gtccccgaga agttgggggg aggggtcggc aattgaacgg gtgcctagag
     2041 aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc tttttcccga
     2101 gggtggggga gaaccgtata taagtgcagt agtcgccgtg aacgttcttt ttcgcaacgg
     2161 gtttgccgcc agaacacagc tgaagcttcg aggggctcgc atctctcctt cacgcgcccg
     2221 ccgccctacc tgaggccgcc atccacgccg gttgagtcgc gttctgccgc ctcccgcctg
     2281 tggtgcctcc tgaactgcgt ccgccgtcta ggtaagttta aagctcaggt cgagaccggg
     2341 cctttgtccg gcgctccctt ggagcctacc tagactcagc cggctctcca cgctttgcct
     2401 gaccctgctt gctcaactct acgtctttgt ttcgttttct gttctgcgcc gttacagatc
     2461 caagctgtga ccggcgccta ggatccatta ccatgagcga gctgattaag gagaacatgc
     2521 acatgaagct gtacatggag ggcaccgtgg acaaccatca cttcaagtgc acatccgagg
     2581 gcgaaggcaa gccctacgag ggcacccaga ccatgagaat caaggtggtc gagggcggcc
     2641 ctctcccctt cgccttcgac atcctggcta ctagcttcct ctacggcagc aagaccttca
     2701 tcaaccacac ccagggcatc cccgacttct tcaagcagtc cttccctgag ggcttcacat
     2761 gggagagagt caccacatac gaagacgggg gcgtgctgac cgctacccag gacaccagcc
     2821 tccaggacgg ctgcctcatc tacaacgtca agatcagagg ggtgaacttc acatccaacg
     2881 gccctgtgat gcagaagaaa acactcggct gggaggcctt caccgagacg ctgtaccccg
     2941 ctgacggcgg cctggaaggc agaaacgaca tggccctgaa gctcgtgggc gggagccatc
     3001 tgatcgcaaa catcaagacc acatatagat ccaagaaacc cgctaagaac ctcaagatgc
     3061 ctggcgtcta ctatgtggac tacagactgg aaagaatcaa ggaggccaac aacgagacct
     3121 acgtcgagca gcacgaggtg gcagtggcca gatactgcga cctccctagc aaactggggc
     3181 acaagcttaa tgcggccgct gagggcagag gaagtcttct aacatgcggt gacgtggagg
     3241 agaatcccgg cccttccgga atgagccggc tggacaagag caaagtgatc aacagcgccc
     3301 tggaactgct gaacggcgtg ggcatcgagg gcctgaccac aagaaagctg gcccagaaac
     3361 tgggcgtgga acagcccacc ctgtactggc acgtgaagaa caagcgggcc ctgctggacg
     3421 ccctgcccat cgagatgctg gacagacacc acacccacag ctgccccctg gaaggcgaga
     3481 gctggcagga cttcctgcgg aacaacgcca agagctacag atgcgccctg ctgagccaca
     3541 gggatggcgc caaagtgcac ctgggcacca gacccaccga gaagcagtac gagacactgg
     3601 aaaaccagct ggccttcctg tgccagcagg gattcagcct ggaaaacgcc ctgtacgccc
     3661 tgagcgccgt gggccacttt acactgggct gcgtgctgga agaacaggaa caccaggtgg
     3721 ccaaagagga aagagagaca cccaccaccg actccatgcc ccccctgctg aagcaggcca
     3781 tcgagctgtt cgatagacag ggcgccgagc ccgcctttct gttcggactg gaactgatca
     3841 tctgcgggct ggaaaagcag ctgaagtgcg agtctggcgg ccctaccgac gccctggacg
     3901 acttcgatct ggacatgctg cctgccgatg ccctggatga ttttgacctg gatatgctgc
     3961 ccgctgacgc tctggacgat ttcgacctgg acatgctgcc cggataggtc gacaatcaac
     4021 ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta
     4081 cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt
     4141 tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg
     4201 ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg
     4261 gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca
     4321 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
     4381 ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctgtg
     4441 ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag
     4501 cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc
     4561 gccctcagac gagtcggatc tccctttggg ccgcctcccc gcctggtacc tttaagacca
     4621 atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa
     4681 gggctaattc actcccaacg aagataagat ctgctttttg cttgtactgg gtctctctgg
     4741 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
     4801 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
     4861 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag tagtagttca
     4921 tgtcatctta ttattcagta tttataactt gcaaagaaat gaatatcaga gagtgagagg
     4981 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     5041 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     5101 tatcatgtct ggctctagct atcccgcccc taactccgcc catcccgccc ctaactccgc
     5161 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     5221 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     5281 acttttgcag agaccaaatt cgtaatcatg tcatagctgt ttcctgtgtg aaattgttat
     5341 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     5401 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     5461 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     5521 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     5581 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     5641 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     5701 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     5761 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     5821 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     5881 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     5941 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     6001 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     6061 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     6121 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     6181 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     6241 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     6301 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     6361 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     6421 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     6481 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     6541 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     6601 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     6661 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     6721 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     6781 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     6841 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     6901 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     6961 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     7021 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     7081 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     7141 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     7201 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     7261 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     7321 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     7381 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     7441 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
     7501 aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac
     7561 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     7621 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat
     7681 gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat accgcacaga
     7741 tgcgtaagga gaaaataccg catcaggcgc cattcgccat tcaggctgcg caactgttgg
     7801 gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct
     7861 gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg
     7921 gccagtgcca agctg
//
