LOCUS       sequence-364219-        5268 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding a self-cleaving peptides p2a sequence 
            (PhCMV-p2A-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5268)
  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 5268)
  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: pTS1018-Tier1-PhCMV-p2A
FEATURES             Location/Qualifiers
     source          1..5268
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(68..85)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(239..827)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(319..338)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(998..1789,1790..1858))
                     /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     1621..1640
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1859..1963)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        2034..2413
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2414..2617
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2568..2588
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2662..2681
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2662..2680
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2742..2798
                     /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(2853..2870)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2859..3083
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3129..3557
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3216..3235)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3426..3447
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3566..3586)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3571..3900
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3751..3886
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3813..3832
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3967..4761
                     /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(4021..4040)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4631..4650
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4935..5056
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4972..4991)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5026..5045
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5105..5121)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5105..5121)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5118..5140)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5129..5145
                     /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(5153..5159,5160..5177,5178..5183))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5198..5219
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
       61 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
      121 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
      181 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
      241 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
      301 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
      361 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
      421 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
      481 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
      541 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
      601 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
      661 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
      721 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
      781 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
      841 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
      901 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
      961 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     1021 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     1081 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     1141 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     1201 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     1261 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     1321 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     1381 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     1441 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     1501 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     1561 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     1621 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     1681 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     1741 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     1801 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     1861 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     1921 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     1981 tgccacctga cgtcgacgga tcgggagatc tccacgcgtg gtaccctcga gttgacattg
     2041 attattgagt agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
     2101 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     2161 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     2221 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     2281 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     2341 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     2401 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
     2461 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
     2521 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
     2581 taggcgtgta cggtgggagg tctatataag cagagctctc tggctaacta gagaacccac
     2641 tgcttactgg cttatcgaaa ttaatacgac tcactatagg gagacccaag ctgttcgaag
     2701 cggaattcac catgactagt ggtggttctg gtggaagcgg agctactaac ttcagcctgc
     2761 tgaagcaggc tggtgacgtg gaggagaacc ctggacctgc tagcggatcc accggtgtct
     2821 agaaagcttt gaggccggcc tgcagggatc agcctcgact gtgccttcta gttgccagcc
     2881 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     2941 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     3001 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     3061 tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg
     3121 gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     3181 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     3241 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     3301 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg
     3361 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     3421 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt
     3481 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     3541 aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
     3601 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
     3661 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     3721 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     3781 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct
     3841 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     3901 aagctcccgg gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg
     3961 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
     4021 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
     4081 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
     4141 gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
     4201 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
     4261 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
     4321 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
     4381 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
     4441 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg
     4501 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
     4561 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
     4621 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
     4681 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
     4741 cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc
     4801 ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     4861 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     4921 tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     4981 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     5041 tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat
     5101 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     5161 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     5221 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgcc
//
