LOCUS       sequence-364210-        5310 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PhCMV-driven PEST tag for destabilization of 
            proteins (PhCMV-PEST-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5310)
  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 5310)
  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: pTS2379-Tier1-PhCMV-PEST
FEATURES             Location/Qualifiers
     source          1..5310
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        114..493
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        494..697
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     648..668
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     742..761
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        742..760
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(975..992)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    981..1205
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1251..1679
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1338..1357)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1548..1569
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1688..1708)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1693..2022
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1873..2008
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1935..1954
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2089..2883
                     /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(2143..2162)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2753..2772
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3057..3178
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3094..3113)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3148..3167
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3227..3243)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3227..3243)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3240..3262)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3251..3267
                     /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(3275..3281,3282..3299,3300..3305))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3320..3341
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3458..3475)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3629..4217)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3709..3728)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4388..5179,5180..5248))
                     /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     5011..5030
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5249..5310,1..43))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
       61 tgccacctga cgtcgacgga tcgggagatc tccacgcgtg gtaccctcga gttgacattg
      121 attattgagt agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
      181 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
      241 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
      301 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
      361 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
      421 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
      481 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
      541 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
      601 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
      661 taggcgtgta cggtgggagg tctatataag cagagctctc tggctaacta gagaacccac
      721 tgcttactgg cttatcgaaa ttaatacgac tcactatagg gagacccaag ctgttcgaag
      781 cggaattcac catgactagt agccatggct tcccgccgga ggtggaggag caggatgatg
      841 gcacgctgcc catgtcttgt gcccaggaga gcgggatgga ccgtcaccct gcagcctgtg
      901 cttctgctag gatcaatgtg gctagcggat ccaccggtgt ctagaaagct ttgaggccgg
      961 cctgcaggga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     1021 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     1081 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     1141 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     1201 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
     1261 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     1321 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     1381 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     1441 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     1501 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     1561 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     1621 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
     1681 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     1741 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     1801 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     1861 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     1921 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
     1981 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
     2041 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     2101 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     2161 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     2221 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     2281 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     2341 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     2401 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     2461 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     2521 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     2581 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     2641 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     2701 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     2761 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     2821 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     2881 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag
     2941 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
     3001 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     3061 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     3121 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     3181 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     3241 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     3301 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     3361 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     3421 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     3481 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     3541 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     3601 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     3661 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     3721 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     3781 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     3841 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     3901 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     3961 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     4021 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     4081 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     4141 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     4201 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     4261 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     4321 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     4381 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     4441 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     4501 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     4561 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     4621 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     4681 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     4741 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     4801 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     4861 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     4921 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     4981 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     5041 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     5101 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     5161 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     5221 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     5281 atcagggtta ttgtctcatg agcggataca
//
