LOCUS       sequence-364234-        5910 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PhCMV-driven tabacco etch virus protease TEV 
            (PhCMV-TEV-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5910)
  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 5910)
  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: pTS2405-Tier1-PhCMV-TEV
FEATURES             Location/Qualifiers
     source          1..5910
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        104..483
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        484..687
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     638..658
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     732..751
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        732..750
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             803..1510
                     /codon_start=1
                     /product="tobacco etch virus protease (Kapust et al., 
                     2001)"
                     /label=TEV protease
                     /translation="GESLFKGPRDYNPISSTICHLTNESDGHTTSLYGIGFGPFIITNK
                     HLFRRNNGTLLVQSLHGVFKVKNTTTLQQHLIDGRDMIIIRMPKDFPPFPQKLKFREPQ
                     REERICLVTTNFQTKSMSSMVSDTSCTFPSSDGIFWKHWIQTKDGQCGSPLVSTRDGFI
                     VGIHSASNFTNTNNYFTSVPKNFMELLTSQEAQQWVSGWRLNADSVLWGGHKVFMSKPE
                     EPFQPVKEATQLMN"
     primer_bind     complement(1565..1582)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1571..1795
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1841..2269
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1928..1947)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2138..2159
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2278..2298)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2283..2612
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2463..2598
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2525..2544
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2679..3473
                     /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(2733..2752)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3343..3362
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3647..3768
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3684..3703)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3738..3757
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3817..3833)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3817..3833)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3830..3852)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3841..3857
                     /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(3865..3871,3872..3889,3890..3895))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3910..3931
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4048..4065)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4219..4807)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4299..4318)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4978..5769,5770..5838))
                     /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     5601..5620
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5839..5910,1..33))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
       61 cgtcgacgga tcgggagatc tccacgcgtg gtaccctcga gttgacattg attattgagt
      121 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
      181 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
      241 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
      301 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
      361 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
      421 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
      481 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
      541 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
      601 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
      661 cggtgggagg tctatataag cagagctctc tggctaacta gagaacccac tgcttactgg
      721 cttatcgaaa ttaatacgac tcactatagg gagacccaag ctgttcgaag cggaattcac
      781 catgactagt ggtggttctg gtggagagag cttgtttaag gggccgcgtg attacaaccc
      841 gatatcgagc accatttgtc atttgacgaa tgaatctgat gggcacacaa catcgttgta
      901 tggtattgga tttggtccct tcatcattac aaacaagcac ttgtttagaa gaaataatgg
      961 aacactgttg gtccaatcac tacatggtgt attcaaggtc aagaacacca cgactttgca
     1021 acaacacctc attgatggga gggacatgat aattattcgc atgcctaagg atttcccacc
     1081 atttcctcaa aagctgaaat ttagagagcc acaaagggaa gagcgcatat gtcttgtgac
     1141 aaccaacttc caaactaaga gcatgtctag catggtgtca gacacaagtt gcacattccc
     1201 ttcatctgat ggcatattct ggaagcattg gattcaaacc aaggatgggc agtgtggcag
     1261 tccattagta tcaactagag atgggttcat tgttggtata cactcagcat cgaatttcac
     1321 caacacaaac aattatttca caagcgtgcc gaaaaacttc atggaattgt tgacaagtca
     1381 ggaggcgcag cagtgggtta gtggttggcg attaaatgct gactcagtat tgtggggggg
     1441 ccataaagtt ttcatgagca aacctgaaga gccttttcag ccagttaagg aagcgactca
     1501 actcatgaat gctagcggat ccaccggtgt ctagaaagct ttgaggccgg cctgcaggga
     1561 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     1621 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     1681 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
     1741 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
     1801 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
     1861 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     1921 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     1981 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     2041 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     2101 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     2161 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     2221 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     2281 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     2341 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     2401 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     2461 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     2521 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     2581 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
     2641 tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     2701 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     2761 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     2821 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     2881 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     2941 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     3001 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     3061 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     3121 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     3181 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     3241 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     3301 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     3361 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     3421 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     3481 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     3541 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     3601 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     3661 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     3721 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     3781 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     3841 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     3901 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     3961 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     4021 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     4081 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     4141 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     4201 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     4261 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     4321 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     4381 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     4441 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     4501 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     4561 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     4621 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     4681 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     4741 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     4801 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     4861 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     4921 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     4981 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     5041 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     5101 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     5161 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     5221 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     5281 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     5341 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     5401 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     5461 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     5521 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     5581 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     5641 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     5701 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     5761 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     5821 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     5881 ttgtctcatg agcggataca tatttgaatg
//
