LOCUS       sequence-364554-        7297 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PVanO2-driven SEAP-p2A-iRFP670 expression 
            (OVanO2-PCMVmin-2-SEAP-p2A-iRFP670-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7297)
  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 7297)
  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: pVH269-Tier1-OVanO2-PCMVmin2-SEAP-p2A-iRFP670
FEATURES             Location/Qualifiers
     source          1..7297
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(88..108)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        93..422
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      273..408
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     335..354
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             489..1283
                     /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(543..562)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1153..1172
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1457..1578
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1494..1513)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1548..1567
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1627..1643)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1627..1643)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1640..1662)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1651..1667
                     /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(1675..1681,1682..1699,1700..1705))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1720..1741
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1858..1875)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2029..2617)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2109..2128)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2788..3579,3580..3648))
                     /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     3411..3430
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3649..3753)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3900..3924
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     3976..3995
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3976..3994
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5625..5681
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             5688..6617
                     /codon_start=1
                     /product="near-infrared fluorescent protein with an 
                     emission peak at 670 nm, engineered from a bacterial 
                     phytochrome (Shcherbakova and Verkhusha, 2013)"
                     /label=iRFP670
                     /note="derived from Rhodopseudomonas palustris RpBphP6"
                     /translation="ARKVDLTSCDREPIHIPGSIQPCGCLLACDAQAVRITRITENAGA
                     FFGRETPRVGELLADYFGETEAHALRNALAQSSDPKRPALIFGWRDGLTGRTFDISLHR
                     HDGTSIIEFEPAAAEQADNPLRLTRQIIARTKELKSLEEMAARVPRYLQAMLGYHRVML
                     YRFADDGSGMVIGEAKRSDLESFLGQHFPASLVPQQARLLYLKNAIRVVSDSRGISSRI
                     VPEHDASGAALDLSFAHLRSISPCHLEFLRNMGVSASMSLSIIIDGTLWGLIICHHYEP
                     RAVPMAQRVAAEMFADFLSLHFTAAHHQR"
     primer_bind     complement(6672..6689)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6678..6902
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      join(6948..7297,1..79)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7035..7054)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7245..7266
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
       61 caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc
      121 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
      181 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
      241 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
      301 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
      361 ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
      421 aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca ggatgaggat
      481 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
      541 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
      601 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
      661 atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
      721 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
      781 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
      841 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
      901 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
      961 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca
     1021 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
     1081 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     1141 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     1201 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     1261 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
     1321 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     1381 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     1441 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     1501 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     1561 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     1621 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     1681 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     1741 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     1801 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     1861 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     1921 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     1981 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     2041 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     2101 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     2161 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     2221 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     2281 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     2341 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     2401 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     2461 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     2521 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     2581 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     2641 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     2701 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     2761 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     2821 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     2881 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     2941 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     3001 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     3061 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     3121 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     3181 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     3241 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     3301 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     3361 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     3421 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     3481 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     3541 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     3601 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     3661 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     3721 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     3781 cgtcgacgga tcgggagatc tccacgcgtg gtaccattgg atccaatagc gctattggat
     3841 ccaatgatac tctcgagtat gtcgaggtgg cgtgtacggt gggaggccta tataagcaga
     3901 gctcgtttag tgaaccgtca gatcgcgagc tctctggcta actagagaac ccactgctta
     3961 ctggcttatc gaaattaata cgactcacta tagggagacc caagctgttc gaagcggaat
     4021 tcaccatgac tagttggctg tggctggtgc tgctgctgct gtgcctggcc ggcaacgtgc
     4081 aggccgctag tatcatcccc gtggaggagg agaaccccga cttctggaac agagaggccg
     4141 ccgaggccct gggcgccgcc aagaagctgc agcccgccca gaccgccgcc aagaacctga
     4201 tcatcttcct gggcgacggc atgggcgtga gcaccgtgac cgccgccaga atcctgaagg
     4261 gccagaagaa ggacaagctg ggccccgaga tccccctggc catggacaga ttcccctacg
     4321 tggccctgag caagacctac aacgtggaca agcacgtgcc cgacagcggc gccaccgcca
     4381 ccgcctacct gtgcggcgtg aagggcaact tccagaccat cggcctgagc gccgccgcca
     4441 gattcaacca gtgcaacacc accagaggca acgaggtgat cagcgtgatg aacagagcca
     4501 agaaggccgg caagagcgtg ggcgtggtga ccaccaccag agtgcagcac gccagccccg
     4561 ccggcaccta cgcccacacc gtgaacagaa actggtacag cgacgccgac gtgcccgcca
     4621 gcgccagaca ggagggctgc caggacatcg ccacccagct gatcagcaac atggacatcg
     4681 acgtgatcct gggcggcggc agaaagtaca tgttcagaat gggcaccccc gaccccgagt
     4741 accccgacga ctacagccag ggcggcacca gactggacgg caagaacctg gtgcaggagt
     4801 ggctggccaa gagacagggc gccagatacg tgtggaacag aaccgagctg atgcaggcca
     4861 gcctggaccc cagcgtgacc cacctgatgg gcctgttcga gcccggcgac atgaagtacg
     4921 agatccacag agacagcacc ctggacccca gcctgatgga gatgaccgag gccgccctga
     4981 gactgctgag cagaaacccc agaggcttct tcctgttcgt ggagggcggc agaatcgacc
     5041 acggccacca cgagagcaga gcctacagag ccctgaccga gaccatcatg ttcgacgacg
     5101 ccatcgagag agccggccag ctgaccagcg aggaggacac cctgagcctg gtgaccgccg
     5161 accacagcca cgtgttcagc ttcggcggct accccctgag aggcagcagc atcttcggcc
     5221 tggcccccgg caaggccaga gacagaaagg cctacaccgt gctgctgtac ggcaacggcc
     5281 ccggctacgt gctgaaggac ggcgccagac ccgacgtgac cgagagcgag agcggcagcc
     5341 ccgagtacag acagcagagc gccgtgcccc tggacgagga gacccacgcc ggcgaggacg
     5401 tggccgtgtt cgccagaggc ccccaggccc acctggtgca cggcgtgcag gagcagacct
     5461 tcatcgccca cgtgatggcc ttcgccgcct gcctggagcc ctacaccgcc tgcgacctgg
     5521 ccccccccgc cggcaccacc gacgccgccc accccggcta cagcagagtg ggcgccgccg
     5581 gcagattcga gcagaccgct agtggtggtt ctggtggaag cggagctact aacttcagcc
     5641 tgctgaagca ggctggtgac gtggaggaga accctggacc tgctagtgcg cgtaaggtcg
     5701 atctcacctc ctgcgatcgc gagccgatcc acatccccgg cagcattcag ccgtgcggct
     5761 gcctgcttgc ctgcgacgcg caggcggtgc ggatcacgcg cattacggaa aatgccggcg
     5821 cgttctttgg acgcgaaact ccgcgggtcg gtgagctact cgccgattac ttcggcgaga
     5881 ccgaagccca tgcgctgcgc aacgcactgg cgcagtcctc cgatccaaag cgaccggcgc
     5941 tgatcttcgg ttggcgcgac ggcctgaccg gccgcacctt cgacatctca ctgcatcgcc
     6001 atgacggtac atcgatcatc gagttcgagc ctgcggcggc cgaacaggcc gacaatccgc
     6061 tgcggctgac gcggcagatc atcgcgcgca ccaaagaact gaagtcgctc gaagagatgg
     6121 ccgcacgggt gccgcgctat ctgcaggcga tgctcggcta tcaccgcgtg atgttgtacc
     6181 gcttcgcgga cgacggctcc gggatggtga tcggcgaggc gaagcgcagc gacctcgaga
     6241 gctttctcgg tcagcacttt ccggcgtcgc tggtcccgca gcaggcgcgg ctactgtact
     6301 tgaagaacgc gatccgcgtg gtctcggatt cgcgcggcat cagcagccgg atcgtgcccg
     6361 agcacgacgc ctccggcgcc gcgctcgatc tgtcgttcgc gcacctgcgc agcatctcgc
     6421 cctgccatct cgaatttctg cggaacatgg gcgtcagcgc ctcgatgtcg ctgtcgatca
     6481 tcattgacgg cacgctatgg ggattgatca tctgtcatca ttacgagccg cgtgccgtgc
     6541 cgatggcgca gcgcgtcgcg gccgaaatgt tcgccgactt cttatcgctg cacttcaccg
     6601 ccgcccacca ccaacgcgct agcggatcca ccggtgtcta gaaagctttg aggccggcct
     6661 gcagggatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     6721 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     6781 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     6841 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     6901 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     6961 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     7021 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     7081 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     7141 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     7201 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     7261 aactggaaca acactcaacc ctatctcggt ctattct
//
