LOCUS       sequence-365223-        5820 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PhCMV-driven tetracycline dependent regulator
            protein TetR (PhCMV-TetR-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5820)
  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 5820)
  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: pTS2365-Tier1-PhCMV-TetR
FEATURES             Location/Qualifiers
     source          1..5820
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        85..464
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        465..668
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     619..639
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     713..732
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        713..731
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             784..1401
                     /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(854..873)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(1456..1473)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1462..1686
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1732..2160
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1819..1838)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2029..2050
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2169..2189)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2174..2503
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2354..2489
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2416..2435
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2570..3364
                     /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(2624..2643)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3234..3253
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3538..3659
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3575..3594)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3629..3648
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3708..3724)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3708..3724)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3721..3743)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3732..3748
                     /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(3756..3762,3763..3780,3781..3786))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3801..3822
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3939..3956)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4110..4698)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4190..4209)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4869..5660,5661..5729))
                     /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     5492..5511
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5730..5820,1..14))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat
       61 ctccacgcgt ggtaccctcg agttgacatt gattattgag tagttattaa tagtaatcaa
      121 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
      181 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
      241 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
      301 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
      361 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
      421 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
      481 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
      541 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta
      601 acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
      661 gcagagctct ctggctaact agagaaccca ctgcttactg gcttatcgaa attaatacga
      721 ctcactatag ggagacccaa gctgttcgaa gcggaattca ccatgactag tggtggttct
      781 ggttccagat tagataaaag taaagtgatt aacagcgcat tagagctgct taatgaggtc
      841 ggaatcgaag gtttaacaac ccgtaaactc gcccagaagc taggtgtaga gcagcctaca
      901 ttgtattggc atgtaaaaaa taagcgggct ttgctcgacg ccttagccat tgagatgtta
      961 gataggcacc atactcactt ttgcccttta gaaggggaaa gctggcaaga ttttttacgt
     1021 aataacgcta aaagttttag atgtgcttta ctaagtcatc gcgatggagc aaaagtacat
     1081 ttaggtacac ggcctacaga aaaacagtat gaaactctcg aaaatcaatt agccttttta
     1141 tgccaacaag gtttttcact agagaatgca ttatatgcac tcagcgctgt ggggcatttt
     1201 actttaggtt gcgtattgga agatcaagag catcaagtcg ctaaagaaga aagggaaaca
     1261 cctactactg atagtatgcc gccattatta cgacaagcta tcgaattatt tgatcaccaa
     1321 ggtgcagagc cagccttctt attcggcctt gaattgatca tatgcggatt agaaaaacaa
     1381 cttaaatgtg aaagtgggtc cgctagcgga tccaccggtg tctagaaagc tttgaggccg
     1441 gcctgcaggg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     1501 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     1561 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     1621 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct
     1681 ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct
     1741 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     1801 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     1861 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     1921 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     1981 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     2041 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     2101 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     2161 aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     2221 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     2281 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     2341 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     2401 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca
     2461 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg
     2521 tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca
     2581 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     2641 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     2701 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc
     2761 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     2821 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     2881 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     2941 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     3001 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     3061 gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct
     3121 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     3181 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     3241 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     3301 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     3361 ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga
     3421 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     3481 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     3541 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     3601 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     3661 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     3721 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     3781 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     3841 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     3901 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     3961 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     4021 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     4081 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     4141 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4201 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4261 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4321 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4381 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4441 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4501 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     4561 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     4621 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     4681 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     4741 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     4801 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     4861 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     4921 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     4981 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     5041 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     5101 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     5161 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     5221 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     5281 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     5341 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     5401 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     5461 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     5521 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     5581 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     5641 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     5701 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     5761 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
//
