LOCUS       sequence-364977-        6277 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PTtgO2-driven SEAP expression 
            (OTtgO2-PCMVmin-2-SEAP-pA). .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6277)
  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 6277)
  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: pVH229-Tier1-OTtgO2-PCMVmin2-SEAP
FEATURES             Location/Qualifiers
     source          1..6277
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(54..70)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(54..70)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(67..89)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    78..94
                     /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(102..108,109..126,127..132))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    147..168
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(285..302)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(456..1044)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(536..555)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1215..2006,2007..2075))
                     /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     1838..1857
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2076..2180)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2327..2351
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     2403..2422
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2403..2421
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4079..4096)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4085..4309
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4355..4783
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4442..4461)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4652..4673
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4792..4812)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4797..5126
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4977..5112
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5039..5058
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5193..5987
                     /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(5247..5266)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5857..5876
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    join(6161..6277,1..5)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6198..6217)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6252..6271
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag
       61 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
      121 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
      181 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
      241 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
      301 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
      361 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
      421 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
      481 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
      541 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
      601 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
      661 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
      721 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
      781 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
      841 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
      901 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
      961 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     1021 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     1081 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     1141 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     1201 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     1261 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     1321 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     1381 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     1441 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     1501 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     1561 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     1621 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     1681 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     1741 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     1801 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     1861 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     1921 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     1981 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     2041 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     2101 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     2161 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt cgacggatcg
     2221 ggagatctcc acgcgtggta ccgtatttac aaacaaccat gaatgtaagt atcatactct
     2281 cgagtatgtc gaggtggcgt gtacggtggg aggcctatat aagcagagct cgtttagtga
     2341 accgtcagat cgcgagctct ctggctaact agagaaccca ctgcttactg gcttatcgaa
     2401 attaatacga ctcactatag ggagacccaa gctgttcgaa gcggaattca ccatgactag
     2461 ttggctgtgg ctggtgctgc tgctgctgtg cctggccggc aacgtgcagg ccgctagtat
     2521 catccccgtg gaggaggaga accccgactt ctggaacaga gaggccgccg aggccctggg
     2581 cgccgccaag aagctgcagc ccgcccagac cgccgccaag aacctgatca tcttcctggg
     2641 cgacggcatg ggcgtgagca ccgtgaccgc cgccagaatc ctgaagggcc agaagaagga
     2701 caagctgggc cccgagatcc ccctggccat ggacagattc ccctacgtgg ccctgagcaa
     2761 gacctacaac gtggacaagc acgtgcccga cagcggcgcc accgccaccg cctacctgtg
     2821 cggcgtgaag ggcaacttcc agaccatcgg cctgagcgcc gccgccagat tcaaccagtg
     2881 caacaccacc agaggcaacg aggtgatcag cgtgatgaac agagccaaga aggccggcaa
     2941 gagcgtgggc gtggtgacca ccaccagagt gcagcacgcc agccccgccg gcacctacgc
     3001 ccacaccgtg aacagaaact ggtacagcga cgccgacgtg cccgccagcg ccagacagga
     3061 gggctgccag gacatcgcca cccagctgat cagcaacatg gacatcgacg tgatcctggg
     3121 cggcggcaga aagtacatgt tcagaatggg cacccccgac cccgagtacc ccgacgacta
     3181 cagccagggc ggcaccagac tggacggcaa gaacctggtg caggagtggc tggccaagag
     3241 acagggcgcc agatacgtgt ggaacagaac cgagctgatg caggccagcc tggaccccag
     3301 cgtgacccac ctgatgggcc tgttcgagcc cggcgacatg aagtacgaga tccacagaga
     3361 cagcaccctg gaccccagcc tgatggagat gaccgaggcc gccctgagac tgctgagcag
     3421 aaaccccaga ggcttcttcc tgttcgtgga gggcggcaga atcgaccacg gccaccacga
     3481 gagcagagcc tacagagccc tgaccgagac catcatgttc gacgacgcca tcgagagagc
     3541 cggccagctg accagcgagg aggacaccct gagcctggtg accgccgacc acagccacgt
     3601 gttcagcttc ggcggctacc ccctgagagg cagcagcatc ttcggcctgg cccccggcaa
     3661 ggccagagac agaaaggcct acaccgtgct gctgtacggc aacggccccg gctacgtgct
     3721 gaaggacggc gccagacccg acgtgaccga gagcgagagc ggcagccccg agtacagaca
     3781 gcagagcgcc gtgcccctgg acgaggagac ccacgccggc gaggacgtgg ccgtgttcgc
     3841 cagaggcccc caggcccacc tggtgcacgg cgtgcaggag cagaccttca tcgcccacgt
     3901 gatggccttc gccgcctgcc tggagcccta caccgcctgc gacctggccc cccccgccgg
     3961 caccaccgac gccgcccacc ccggctacag cagagtgggc gccgccggca gattcgagca
     4021 gaccgctagc ggatccaccg gtgtctagaa agctttgagg ccggcctgca gggatcagcc
     4081 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     4141 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
     4201 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     4261 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg
     4321 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc
     4381 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     4441 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     4501 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     4561 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     4621 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     4681 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     4741 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt
     4801 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     4861 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     4921 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     4981 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     5041 tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct
     5101 tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat
     5161 ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag
     5221 gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg
     5281 gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca
     5341 agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc
     5401 tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg
     5461 actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg
     5521 ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta
     5581 cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag
     5641 ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac
     5701 tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg
     5761 atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg
     5821 gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg
     5881 aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg
     5941 attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg
     6001 gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc
     6061 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     6121 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     6181 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     6241 gcattctagt tgtggtttgt ccaaactcat caatgta
//
