LOCUS       sequence-364222-        5796 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PhCMV-driven marker for puromycin selection 
            (PhCMV-PuroR-pA).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5796)
  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 5796)
  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: pTS1021-Tier1-PhCMV-PuroR
FEATURES             Location/Qualifiers
     source          1..5796
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        101..480
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        481..684
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     635..655
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     729..748
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        729..747
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             800..1393
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     1293..1313
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(1448..1465)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1454..1678
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1724..2152
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1811..1830)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2021..2042
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2161..2181)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2166..2495
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2346..2481
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2408..2427
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2562..3356
                     /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(2616..2635)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3226..3245
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3530..3651
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3567..3586)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3621..3640
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3700..3716)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3700..3716)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3713..3735)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3724..3740
                     /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(3748..3754,3755..3772,3773..3778))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3793..3814
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3931..3948)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4102..4690)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4182..4201)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4861..5652,5653..5721))
                     /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     5484..5503
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5722..5796,1..30))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
       61 cgacggatcg ggagatctcc acgcgtggta ccctcgagtt gacattgatt attgagtagt
      121 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
      181 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
      241 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
      301 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
      361 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
      421 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
      481 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
      541 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      601 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      661 tgggaggtct atataagcag agctctctgg ctaactagag aacccactgc ttactggctt
      721 atcgaaatta atacgactca ctatagggag acccaagctg ttcgaagcgg aattcaccat
      781 gactagtggt ggttctggta ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga
      841 cgtcccccgg gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca
      901 caccgtcgac ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac
      961 gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt
     1021 ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat
     1081 ggccgagttg agcggttccc ggctggccgc gcagcaacag atggaaggcc tcctggcgcc
     1141 gcaccggccc aaggagcccg cgtggttcct ggccaccgtc ggcgtgtcgc ccgaccacca
     1201 gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg
     1261 ggtgcccgcc ttcctggaga catccgcgcc ccgcaacctc cccttctacg agcggctcgg
     1321 cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg
     1381 caagcccggt gccgctagcg gatccaccgg tgtctagaaa gctttgaggc cggcctgcag
     1441 ggatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     1501 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     1561 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     1621 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
     1681 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
     1741 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     1801 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     1861 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     1921 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     1981 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     2041 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     2101 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt
     2161 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     2221 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     2281 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     2341 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     2401 ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt
     2461 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca
     2521 ttttcggatc tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat
     2581 tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac
     2641 agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc
     2701 tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc
     2761 tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag
     2821 cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc
     2881 ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg
     2941 atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc
     3001 ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc
     3061 cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga
     3121 cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca
     3181 tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg
     3241 atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg
     3301 ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg
     3361 gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga
     3421 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     3481 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     3541 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     3601 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     3661 tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     3721 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     3781 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     3841 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     3901 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     3961 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     4021 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     4081 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     4141 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     4201 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     4261 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     4321 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     4381 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     4441 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     4501 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     4561 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     4621 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     4681 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     4741 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     4801 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     4861 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     4921 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     4981 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     5041 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     5101 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     5161 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     5221 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     5281 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     5341 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     5401 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     5461 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     5521 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     5581 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     5641 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     5701 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     5761 ttattgtctc atgagcggat acatatttga atgtat
//
