LOCUS       sequence-172790-        7487 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7487)
  AUTHORS   Kita-Matsuo H, Barcova M, Prigozhina N, Salomonis N, Wei K, Jacot 
            JG, Nelson B, Spiering S, Haverslag R, Kim C, Talantova M, Bajpai R,
            Calzolari D, Terskikh A, McCulloch AD, Price JH, Conklin BR, Chen 
            HS, Mercola M
  TITLE     Lentiviral vectors and protocols for creation of stable hESC lines 
            for fluorescent tracking and drug resistance selection of 
            cardiomyocytes.
  JOURNAL   PLoS ONE. 2009  . 4(4):e5046.
  PUBMED    19352491
REFERENCE   2  (bases 1 to 7487)
  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: "PLoS ONE. 
            2009  . 4(4):e5046."
COMMENT     Sequence Label: Brachyury-eGFP
FEATURES             Location/Qualifiers
     source          1..7487
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        27..131
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(132..200,201..992)
                     /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     complement(350..369)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1163..1751
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1652..1671
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1905..1922
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2039..2060
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2075..2080,2081..2098,2099..2105)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2113..2129
                     /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)."
     primer_bind     2118..2140
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2137..2153
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2137..2153
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2172..2192
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2174..2192
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     LTR             2448..2628
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    2672..2797
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    3290..3523
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             3708..3752
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    4019..4135
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     regulatory      4812..4821
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4818..4820,4821..4823,4824..5537)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(4818..4820,4821..4823,4824..5534)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4863..4884)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5124..5143)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5471..5492
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    5553..6141
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5606..5626)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6024..6035)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             6228..6461
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    6533..6654
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6570..6589)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6624..6643
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      6673..6808
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6840..6859)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6841..6859)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6869..6886)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6869..6885)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6878..6900)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      7030..7485
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7117..7136)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7327..7348
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
       61 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
      121 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
      181 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
      241 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
      301 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
      361 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
      421 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
      481 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
      541 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
      601 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
      661 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
      721 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
      781 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
      841 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
      901 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
      961 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     1021 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     1081 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     1141 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     1201 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     1261 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     1321 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     1381 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     1441 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     1501 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     1561 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     1621 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     1681 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     1741 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     1801 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     1861 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     1921 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     1981 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     2041 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     2101 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
     2161 acgccaagcg cgcaattaac cctcactaaa gggaacaaaa gctggagctg caagcttaat
     2221 gtagtcttat gcaatactct tgtagtcttg caacatggta acgatgagtt agcaacatgc
     2281 cttacaagga gagaaaaagc accgtgcatg ccgattggtg gaagtaaggt ggtacgatcg
     2341 tgccttatta ggaaggcaac agacgggtct gacatggatt ggacgaacca ctgaattgcc
     2401 gcattgcaga gatattgtat ttaagtgcct agctcgatac aataaacggg tctctctggt
     2461 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
     2521 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
     2581 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa
     2641 cagggacctg aaagcgaaag ggaaaccaga gctctctcga cgcaggactc ggcttgctga
     2701 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag
     2761 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag
     2821 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca
     2881 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac
     2941 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga
     3001 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga
     3061 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac
     3121 caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt
     3181 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
     3241 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
     3301 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcctcaatg acgctgacgg
     3361 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
     3421 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
     3481 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
     3541 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
     3601 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca
     3661 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag
     3721 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc
     3781 tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt
     3841 ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga
     3901 cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag
     3961 agagagacag agacagatcc attcgattag tgaacggatc tcgacggtat cggttaactt
     4021 ttaaaagaaa aggggggatt ggggggtaca gtgcagggga aagaatagta gacataatag
     4081 caacagacat acaaacaaag aattacaaaa acaaattaca aaaattcaaa attttatcga
     4141 tcacgagact agcctcgagg gcaaagtcgc aggcgccggt gtgcgcttgg acagcgcgtg
     4201 ggagtggaga gtttagcagt ggctctagga gccagggtcc tgggtggctc cagcccggct
     4261 tctcgccctc cctcccccag ggtccgcccc gccgctttga tggaggtgca aacatttggg
     4321 ggagggcggg ggtgtcggga ctgcgcccga cgctttcctt acaggaagcg cgcgctggag
     4381 cccattgttg gcccccagcc tccgggcccg cccggccagt ctgatatggc cgcgcaccgc
     4441 caatgggcag ctgctcggta cttcaaaggg tgtcccgccc aatccgccgc acccccctgc
     4501 gaggccacct cggctgtatt tatggggagg ggacccattt ttctcttccc cagagactta
     4561 ctcttgtcgc gccttgcggg agttcaagtg gagccacggc tccccaggcc ctctccccca
     4621 tccccgcccc cttcccccct catcccgatc tcggtgctcc tttggcgaat gtgcagggac
     4681 ccaggtgtaa tctttgggct ccgcagagtg accctttttc ttggaaaagc ggtggcgaga
     4741 gaagtgaagg tggctgttgg gtagggagtc aagactcctg gaaggtggag agggtggcgg
     4801 gaggaccggt cgccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca
     4861 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg
     4921 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc
     4981 ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct
     5041 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc
     5101 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt
     5161 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg
     5221 gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg
     5281 ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg
     5341 gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc
     5401 tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga
     5461 agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg
     5521 acgagctgta caagtaaagc ggccgcgtcg acaatcaacc tctggattac aaaatttgtg
     5581 aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt
     5641 taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata
     5701 aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg
     5761 tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc
     5821 tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct
     5881 gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt
     5941 cggggaagct gacgtccttt ccatggctgc tcgcctgtgt tgccacctgg attctgcgcg
     6001 ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc
     6061 tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct
     6121 ccctttgggc cgcctccccg cctggaattc gagctcggta cctttaagac caatgactta
     6181 caaggcagct gtagatctta gccacttttt aaaagaaaag gggggactgg aagggctaat
     6241 tcactcccaa cgaagacaag atctgctttt tgcttgtact gggtctctct ggttagacca
     6301 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag
     6361 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag
     6421 atccctcaga cccttttagt cagtgtggaa aatctctagc agtagtagtt catgtcatct
     6481 tattattcag tatttataac ttgcaaagaa atgaatatca gagagtgaga ggaacttgtt
     6541 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     6601 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     6661 ctggctctag ctatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     6721 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     6781 aagtagtgag gaggcttttt tggaggccta ggcttttgcg tcgagacgta cccaattcgc
     6841 cctatagtga gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg
     6901 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
     6961 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
     7021 aatggcgcga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     7081 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     7141 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     7201 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     7261 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     7321 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     7381 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     7441 aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aatttcc
//
