LOCUS       sequence-365224-        5952 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PhCMV-driven tetracycline-dependent 
            F-type-based transactiavtor (PhCMV-TetR-F-type-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5952)
  AUTHORS   Haellman V, Strittmatter T, Bertschi A, Stucheli P, Fussenegger M
  TITLE     A versatile plasmid architecture for mammalian synthetic biology 
            (VAMSyB).
  JOURNAL   Metab Eng. 2021 Jul;66:41-50. doi: 10.1016/j.ymben.2021.04.003. Epub
            2021 Apr 18.
  PUBMED    33857582
REFERENCE   2  (bases 1 to 5952)
  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: "Metab 
            Eng."; date: "2021-07"; volume: "66:41-50. doi"; pages: " 
            10.1016/j.ymben.2021.04.003. Epub 2021 Apr 18"
COMMENT     Sequence Label: pTS2370-Tier1-PhCMV-TetR_F-type
FEATURES             Location/Qualifiers
     source          1..5952
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        79..458
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        459..662
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     613..633
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     707..726
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        707..725
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             778..1395
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="SRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHVK
                     NKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTRP
                     TEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTTD
                     SMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(848..867)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     CDS             1411..1521
                     /codon_start=1
                     /product="3 tandem repeats of the minimal activation domain
                     of herpes simplex virus VP16 (Cheng et al., 2013)"
                     /label=VP48
                     /translation="DALDDFDLDMLPADALDDFDLDMLPADALDDFDLDML"
     primer_bind     complement(1582..1599)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1588..1812
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1858..2286
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1945..1964)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2155..2176
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2295..2315)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2300..2629
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2480..2615
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2542..2561
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2696..3490
                     /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"
     primer_bind     complement(2750..2769)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3360..3379
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3664..3785
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3701..3720)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3755..3774
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3834..3850)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3834..3850)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3847..3869)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3858..3874
                     /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(3882..3888,3889..3906,3907..3912))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3927..3948
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4065..4082)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4236..4824)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4316..4335)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4995..5786,5787..5855))
                     /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     5618..5637
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5856..5952,1..8))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctccac
       61 gcgtggtacc ctcgagttga cattgattat tgagtagtta ttaatagtaa tcaattacgg
      121 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
      181 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
      241 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
      301 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
      361 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
      421 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
      481 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
      541 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
      601 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
      661 ctctctggct aactagagaa cccactgctt actggcttat cgaaattaat acgactcact
      721 atagggagac ccaagctgtt cgaagcggaa ttcaccatga ctagtggtgg ttctggttcc
      781 agattagata aaagtaaagt gattaacagc gcattagagc tgcttaatga ggtcggaatc
      841 gaaggtttaa caacccgtaa actcgcccag aagctaggtg tagagcagcc tacattgtat
      901 tggcatgtaa aaaataagcg ggctttgctc gacgccttag ccattgagat gttagatagg
      961 caccatactc acttttgccc tttagaaggg gaaagctggc aagatttttt acgtaataac
     1021 gctaaaagtt ttagatgtgc tttactaagt catcgcgatg gagcaaaagt acatttaggt
     1081 acacggccta cagaaaaaca gtatgaaact ctcgaaaatc aattagcctt tttatgccaa
     1141 caaggttttt cactagagaa tgcattatat gcactcagcg ctgtggggca ttttacttta
     1201 ggttgcgtat tggaagatca agagcatcaa gtcgctaaag aagaaaggga aacacctact
     1261 actgatagta tgccgccatt attacgacaa gctatcgaat tatttgatca ccaaggtgca
     1321 gagccagcct tcttattcgg ccttgaattg atcatatgcg gattagaaaa acaacttaaa
     1381 tgtgaaagtg ggtccgctag tgcaccggcc gacgcccttg acgattttga cttagacatg
     1441 ctcccagccg atgcccttga cgactttgac cttgatatgc tgcctgctga cgctcttgac
     1501 gattttgacc ttgacatgct ccccggggct agcggatcca ccggtgtcta gaaagctttg
     1561 aggccggcct gcagggatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     1621 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     1681 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     1741 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     1801 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
     1861 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     1921 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     1981 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     2041 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     2101 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     2161 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     2221 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     2281 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     2341 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     2401 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     2461 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     2521 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     2581 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     2641 agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat
     2701 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     2761 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     2821 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     2881 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     2941 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     3001 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     3061 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     3121 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     3181 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     3241 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     3301 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     3361 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     3421 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     3481 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     3541 tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     3601 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     3661 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     3721 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     3781 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag
     3841 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     3901 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     3961 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     4021 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4081 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4141 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     4201 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     4261 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     4321 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     4381 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     4441 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     4501 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     4561 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     4621 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     4681 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     4741 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     4801 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     4861 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     4921 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     4981 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5041 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5101 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     5161 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     5221 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     5281 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     5341 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     5401 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     5461 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     5521 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     5581 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     5641 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     5701 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     5761 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     5821 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     5881 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     5941 aaacaaatag gg
//
