LOCUS       sequence-365009-        5346 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PhCMV-driven VP64 transactivator domain 
            (PhCMV-VP64-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5346)
  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 5346)
  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: pTS2368-Tier1-PhCMV_VP64
FEATURES             Location/Qualifiers
     source          1..5346
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        59..438
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        439..642
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     593..613
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     687..706
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        687..705
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             746..895
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     primer_bind     complement(956..973)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    962..1186
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1232..1660
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1319..1338)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1529..1550
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1669..1689)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1674..2003
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1854..1989
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1916..1935
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2070..2864
                     /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(2124..2143)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2734..2753
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3038..3159
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3075..3094)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3129..3148
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3208..3224)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3208..3224)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3221..3243)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3232..3248
                     /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(3256..3262,3263..3280,3281..3286))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3301..3322
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3439..3456)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3610..4198)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3690..3709)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4369..5160,5161..5229))
                     /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     4992..5011
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5230..5334)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaaagtgcca cctgacgtcg acggatcggg agatctccac gcgtggtacc ctcgagttga
       61 cattgattat tgagtagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
      121 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
      181 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
      241 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      301 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      361 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      421 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
      481 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
      541 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
      601 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa
      661 cccactgctt actggcttat cgaaattaat acgactcact atagggagac ccaagctgtt
      721 cgaagcggaa ttcaccatga ctagtgacgc gctggacgat ttcgatctcg acatgctggg
      781 ttctgatgcc ctcgatgact ttgacctgga tatgttggga agcgacgcat tggatgactt
      841 tgatctggac atgctcggct ccgatgctct ggacgatttc gatctcgata tgttaattaa
      901 cgctagcgga tccaccggtg tctagaaagc tttgaggccg gcctgcaggg atcagcctcg
      961 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     1021 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     1081 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     1141 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa
     1201 agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg
     1261 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     1321 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     1381 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     1441 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     1501 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     1561 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     1621 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc
     1681 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     1741 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     1801 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     1861 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     1921 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt
     1981 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg
     2041 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     2101 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     2161 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     2221 ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg
     2281 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     2341 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     2401 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     2461 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     2521 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     2581 tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     2641 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     2701 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     2761 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     2821 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     2881 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     2941 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     3001 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa
     3061 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     3121 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac
     3181 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     3241 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     3301 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     3361 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     3421 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     3481 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     3541 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     3601 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     3661 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     3721 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     3781 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     3841 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     3901 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     3961 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     4021 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     4081 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     4141 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4201 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     4261 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     4321 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     4381 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     4441 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     4501 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     4561 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     4621 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     4681 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     4741 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     4801 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     4861 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     4921 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     4981 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     5041 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     5101 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     5161 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     5221 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     5281 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     5341 tccccg
//
