LOCUS       sequence-364607-        5994 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PhCMV-driven blue fluorescent protein 
            mTagBFP2 fused to a ""self-cleaving"" peptide p2a 
            (PhCMV-p2a_mTagBFP2-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5994)
  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 5994)
  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: pTS2387-Tier1-PhCMV-p2a_mTagBFP2
FEATURES             Location/Qualifiers
     source          1..5994
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        5..334
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      185..320
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     247..266
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             401..1195
                     /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(455..474)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1065..1084
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1369..1490
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1406..1425)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1460..1479
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1539..1555)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1539..1555)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1552..1574)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1563..1579
                     /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(1587..1593,1594..1611,1612..1617))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1632..1653
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1770..1787)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1941..2529)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2021..2040)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2700..3491,3492..3560))
                     /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     3323..3342
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3561..3665)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        3736..4115
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4116..4319
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4270..4290
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4364..4383
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4364..4382
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4444..4500
                     /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"
     CDS             4519..5226
                     /codon_start=1
                     /product="enhanced monomeric blue fluorescent protein 
                     (Subach et al., 2011)"
                     /label=mTagBFP2
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMR
                     IKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGV
                     LTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDM
                     ALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVA
                     RYCDLPSKLGHKLN"
     primer_bind     complement(5281..5298)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5287..5511
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5557..5985
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5644..5663)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5854..5875
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(join(5994,1..20))
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
ORIGIN
        1 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
       61 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
      121 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
      181 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
      241 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
      301 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
      361 tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
      421 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
      481 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
      541 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
      601 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
      661 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
      721 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
      781 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
      841 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
      901 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
      961 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     1021 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     1081 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     1141 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     1201 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     1261 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     1321 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
     1381 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     1441 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     1501 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
     1561 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     1621 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     1681 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     1741 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     1801 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     1861 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     1921 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     1981 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     2041 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     2101 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     2161 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     2221 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     2281 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     2341 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     2401 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     2461 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     2521 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     2581 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     2641 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     2701 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     2761 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     2821 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     2881 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     2941 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     3001 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     3061 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     3121 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     3181 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     3241 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     3301 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     3361 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     3421 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     3481 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     3541 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     3601 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     3661 ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctccacgcg
     3721 tggtaccctc gagttgacat tgattattga gtagttatta atagtaatca attacggggt
     3781 cattagttca tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc
     3841 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag
     3901 taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc
     3961 acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg
     4021 gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc
     4081 agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca
     4141 atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca
     4201 atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg
     4261 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctc
     4321 tctggctaac tagagaaccc actgcttact ggcttatcga aattaatacg actcactata
     4381 gggagaccca agctgttcga agcggaattc accatgacta gtggtggttc tggtggaagc
     4441 ggagctacta acttcagcct gctgaagcag gctggtgacg tggaggagaa ccctggacct
     4501 gctagtggtg gttctggtgt gtctaagggc gaagagctga ttaaggagaa catgcacatg
     4561 aagctgtaca tggagggcac cgtggacaac catcacttca agtgcacatc cgagggcgaa
     4621 ggcaagccct acgagggcac ccagaccatg agaatcaagg tggtcgaggg cggccctctc
     4681 cccttcgcct tcgacatcct ggctactagc ttcctctacg gcagcaagac cttcatcaac
     4741 cacacccagg gcatccccga cttcttcaag cagtccttcc ctgagggctt cacatgggag
     4801 agagtcacca catacgaaga cgggggcgtg ctgaccgcta cccaggacac cagcctccag
     4861 gacggctgcc tcatctacaa cgtcaagatc agaggggtga acttcacatc caacggccct
     4921 gtgatgcaga agaaaacact cggctgggag gccttcaccg agacgctgta ccccgctgac
     4981 ggcggcctgg aaggcagaaa cgacatggcc ctgaagctcg tgggcgggag ccatctgatc
     5041 gcaaacgcca agaccacata tagatccaag aaacccgcta agaacctcaa gatgcctggc
     5101 gtctactatg tggactacag actggaaaga atcaaggagg ccaacaacga gacctacgtc
     5161 gagcagcacg aggtggcagt ggccagatac tgcgacctcc ctagcaaact ggggcacaaa
     5221 cttaatgcta gcggatccac cggtgtctag aaagctttga ggccggcctg cagggatcag
     5281 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     5341 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     5401 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     5461 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg
     5521 cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa
     5581 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     5641 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     5701 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     5761 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     5821 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     5881 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     5941 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtg
//
