LOCUS       sequence-366598-        5804 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector carrying an internal ribosome entry site (IRES-pA).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5804)
  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 5804)
  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: pTS2364-Tier1-IRES
FEATURES             Location/Qualifiers
     source          1..5804
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        33..412
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        413..616
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     567..587
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     661..680
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        661..679
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(744..1318,1319..1321,1322..1330)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(911..928)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1138..1157
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     complement(1388..1405)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1394..1618
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1664..2092
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1751..1770)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1961..1982
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2101..2121)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2106..2435
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2286..2421
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2348..2367
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2502..3296
                     /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(2556..2575)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3166..3185
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3470..3591
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3507..3526)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3561..3580
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3640..3656)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3640..3656)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3653..3675)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3664..3680
                     /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(3688..3694,3695..3712,3713..3718))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3733..3754
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3871..3888)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4042..4630)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4122..4141)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4801..5592,5593..5661))
                     /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     5424..5443
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5662..5766)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgggagatct ccacgcgtgg taccctcgag ttgacattga ttattgagta gttattaata
       61 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
      121 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
      181 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
      241 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc
      301 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg
      361 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
      421 gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
      481 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
      541 atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt
      601 ctatataagc agagctctct ggctaactag agaacccact gcttactggc ttatcgaaat
      661 taatacgact cactataggg agacccaagc tgttcgaagc ggaattcacc atgactagtt
      721 agaaccgttc cgagtcgtta acgcccctct ccctcccccc cccctaacgt tactggccga
      781 agccgcttgg aataaggccg gtgtgcgttt gtctatatgt tattttccac catattgccg
      841 tcttttggca atgtgagggc ccggaaacct ggccctgtct tcttgacgag cattcctagg
      901 ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa ggaagcagtt
      961 cctctggaag cctcttgaag acaaacaacg tctgtagcga ccctttgcag gcagcggaac
     1021 cccccacctg gcgacaggtg cctctgcggc caaaagccac gtgtataaga tacacctgca
     1081 aaggcggcac aaccccagtg ccacgttgtg agttggatag ttgtggaaag agtcaaatgg
     1141 ctctcctcaa gcgtattcaa caaggggctg aaggatgccc agaaggtacc ccattgtatg
     1201 ggatctgatc tggggcctcg gtacacatgc tttacatgtg tttagtcgag gttaaaaaaa
     1261 cgtctaggcc ccccgaacca cggggacgtg gttttccttt gaaaaacacg atgataaact
     1321 tgccacaacc atggctagcg gatccaccgg tgtctagaaa gctttgaggc cggcctgcag
     1381 ggatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     1441 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     1501 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     1561 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
     1621 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
     1681 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     1741 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     1801 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     1861 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     1921 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     1981 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     2041 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt
     2101 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     2161 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     2221 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     2281 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     2341 ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt
     2401 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca
     2461 ttttcggatc tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat
     2521 tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac
     2581 agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc
     2641 tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc
     2701 tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag
     2761 cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc
     2821 ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg
     2881 atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc
     2941 ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc
     3001 cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga
     3061 cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca
     3121 tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg
     3181 atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg
     3241 ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg
     3301 gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga
     3361 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     3421 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     3481 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     3541 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     3601 tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     3661 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     3721 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     3781 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     3841 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     3901 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     3961 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     4021 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     4081 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     4141 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     4201 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     4261 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     4321 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     4381 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     4441 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     4501 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     4561 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     4621 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     4681 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     4741 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     4801 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     4861 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     4921 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     4981 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     5041 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     5101 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     5161 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     5221 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     5281 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     5341 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     5401 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     5461 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     5521 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     5581 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     5641 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     5701 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     5761 tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggat
//
