LOCUS       sequence-364992-        7763 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PTK-driven Citrine-p2A-SEAP expression 
            (PTK-Citrine-p2A-SEAP-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7763)
  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 7763)
  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: pVH65-Tier1-PTK-Citrine_p2a_SEAP
FEATURES             Location/Qualifiers
     source          1..7763
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      167..302
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     229..248
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             383..1177
                     /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(437..456)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1047..1066
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1351..1472
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1388..1407)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1442..1461
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1521..1537)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1521..1537)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1534..1556)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1545..1561
                     /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(1569..1575,1576..1593,1594..1599))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1614..1635
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1752..1769)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1923..2511)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2003..2022)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2682..3473,3474..3542))
                     /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     3305..3324
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3543..3647)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3717..4468
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     primer_bind     4516..4535
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4516..4534
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4890..4909)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             5322..5378
                     /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"
     primer_bind     complement(7032..7049)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7038..7262
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      7308..7736
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7395..7414)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7605..7626
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(join(7745..7763,1..2))
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        join(7750..7763,1..316)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
ORIGIN
        1 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
       61 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
      121 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
      181 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
      241 ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
      301 cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcaaga
      361 gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc
      421 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
      481 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
      541 gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac
      601 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
      661 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
      721 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
      781 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
      841 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
      901 gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
      961 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     1021 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     1081 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     1141 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg
     1201 accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat
     1261 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     1321 gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac
     1381 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     1441 tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc
     1501 tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     1561 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     1621 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     1681 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     1741 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     1801 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     1861 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     1921 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     1981 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     2041 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     2101 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     2161 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     2221 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     2281 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     2341 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     2401 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     2461 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     2521 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     2581 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     2641 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     2701 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     2761 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     2821 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     2881 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     2941 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     3001 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     3061 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     3121 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     3181 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     3241 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     3301 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     3361 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     3421 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     3481 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     3541 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     3601 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     3661 aaagtgccac ctgacgtcga cggatcggga gatctccacg cgtggtaccc tcgagaaatg
     3721 agtcttcgga cctcgcgggg gccgcttaag cggtggttag ggtttgtctg acgcgggggg
     3781 agggggaagg aacgaaacac tctcattcgg aggcggctcg gggtttggtc ttggtggcca
     3841 cgggcacgca gaagagcgcc gcgatcctct taagcacccc cccgccctcc gtggaggcgg
     3901 gggtttggtc ggcgggtggt aactggcggg ccgctgactc gggcgggtcg cgcgccccag
     3961 agtgtgacct tttcggtctg ctcgcagacc cccgggcggc gccgccgcgg cggcgacggg
     4021 ctcgctgggt cctaggctcc atggggaccg tatacgtgga caggctctgg agcatccgca
     4081 cgactgcggt gatattaccg gagaccttct gcgggacgag ccgggtcacg cggctgacgc
     4141 ggagcgtccg ttgggcgaca aacaccagga cggggcacag gtacactatc ttgtcacccg
     4201 gaggcgcgag ggactgcagg agcttcaggg agtggcgcag ctgcttcatc cccgtggccc
     4261 gttgctcgcg tttgctggcg gtgtccccgg aagaaatata tttgcatgtc tttagttcta
     4321 tgatgacaca aaccccgccc agcgtcttgt cattggcgaa ttcgaacacg cagatgcagt
     4381 cggggcggcg cggtcccagg tccacttcgc atattaaggt gacgcgtgtg gcctcgaaca
     4441 ccgagcgacc ctgcagcgac ccgcttaaga gctctggcta actagagaac ccactgctta
     4501 ctggcttatc gaaattaata cgactcacta tagggagacc caagctgttc gaagcggaat
     4561 tcaccatgac tagtggtggt tctggtgtga gcaagggcga ggagctgttc accggcgtgg
     4621 tgcccatcct ggtggagctg gacggcgacg tgaacggcca caagttcagc gtgagcggcg
     4681 agggcgaggg cgacgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca
     4741 agctgcccgt gccctggccc accctggtga ccaccttcgg ctacggcctg atgtgcttcg
     4801 ccagataccc cgaccacatg aagcagcacg acttcttcaa gagcgccatg cccgagggct
     4861 acgtgcagga gagaaccatc ttcttcaagg acgacggcaa ctacaagacc agagccgagg
     4921 tgaagttcga gggcgacacc ctggtgaaca gaatcgagct gaagggcatc gacttcaagg
     4981 aggacggcaa catcctgggc cacaagctgg agtacaacta caacagccac aacgtgtaca
     5041 tcatggccga caagcagaag aacggcatca aggtgaactt caagatcaga cacaacatcg
     5101 aggacggcag cgtgcagctg gccgaccact accagcagaa cacccccatc ggcgacggcc
     5161 ccgtgctgct gcccgacaac cactacctga gctaccagag cgccctgagc aaggacccca
     5221 acgagaagag agaccacatg gtgctgctgg agttcgtgac cgccgccggc atcaccctgg
     5281 gcatggacga gctgtacaag accggtcaca tgggcagcgg cgccaccaac ttcagcctgc
     5341 tgaagcaggc cggcgacgtg gaggagaacc ccggccccga cgtgcccccc tgccagagcc
     5401 agagaagcag agactctagt atgctgctgc tgctgctgct gctgggcctg agactgcagc
     5461 tgagcctggg catcatcccc gtggaggagg agaaccccga cttctggaac agagaggccg
     5521 ccgaggccct gggcgccgcc aagaagctgc agcccgccca gaccgccgcc aagaacctga
     5581 tcatcttcct gggcgacggc atgggcgtga gcaccgtgac cgccgccaga atcctgaagg
     5641 gccagaagaa ggacaagctg ggccccgaga tccccctggc catggacaga ttcccctacg
     5701 tggccctgag caagacctac aacgtggaca agcacgtgcc cgacagcggc gccaccgcca
     5761 ccgcctacct gtgcggcgtg aagggcaact tccagaccat cggcctgagc gccgccgcca
     5821 gattcaacca gtgcaacacc accagaggca acgaggtgat cagcgtgatg aacagagcca
     5881 agaaggccgg caagagcgtg ggcgtggtga ccaccaccag agtgcagcac gccagccccg
     5941 ccggcaccta cgcccacacc gtgaacagaa actggtacag cgacgccgac gtgcccgcca
     6001 gcgccagaca ggagggctgc caggacatcg ccacccagct gatcagcaac atggacatcg
     6061 acgtgatcct gggcggcggc agaaagtaca tgttcagaat gggcaccccc gaccccgagt
     6121 accccgacga ctacagccag ggcggcacca gactggacgg caagaacctg gtgcaggagt
     6181 ggctggccaa gagacagggc gccagatacg tgtggaacag aaccgagctg atgcaggcca
     6241 gcctggaccc cagcgtgacc cacctgatgg gcctgttcga gcccggcgac atgaagtacg
     6301 agatccacag agacagcacc ctggacccca gcctgatgga gatgaccgag gccgccctga
     6361 gactgctgag cagaaacccc agaggcttct tcctgttcgt ggagggcggc agaatcgacc
     6421 acggccacca cgagagcaga gcctacagag ccctgaccga gaccatcatg ttcgacgacg
     6481 ccatcgagag agccggccag ctgaccagcg aggaggacac cctgagcctg gtgaccgccg
     6541 accacagcca cgtgttcagc ttcggcggct accccctgag aggcagcagc atcttcggcc
     6601 tggcccccgg caaggccaga gacagaaagg cctacaccgt gctgctgtac ggcaacggcc
     6661 ccggctacgt gctgaaggac ggcgccagac ccgacgtgac cgagagcgag agcggcagcc
     6721 ccgagtacag acagcagagc gccgtgcccc tggacgagga gacccacgcc ggcgaggacg
     6781 tggccgtgtt cgccagaggc ccccaggccc acctggtgca cggcgtgcag gagcagacct
     6841 tcatcgccca cgtgatggcc ttcgccgcct gcctggagcc ctacaccgcc tgcgacctgg
     6901 ccccccccgc cggcaccacc gacgccgccc accccggcta cagcagagtg ggcgccgccg
     6961 gcagattcga gcagaccgct agcggatcca ccggtgtcta gaaagctttg aggccggcct
     7021 gcagggatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     7081 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     7141 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     7201 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     7261 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     7321 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     7381 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     7441 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     7501 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     7561 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     7621 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     7681 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
     7741 ctgtggaatg tgtgtcagtt agg
//
