LOCUS       sequence-357225-        8984 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  for lentiviral expression of brachyury ORF in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8984)
  TITLE     brachyury ORF lentiviral expression plasmids
REFERENCE   2  (bases 1 to 8984)
  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"
COMMENT     Sequence Label: Phage-PGK-FLAG-HA-T(WT)
FEATURES             Location/Qualifiers
     source          1..8984
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..634
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1721..1765
                     /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    2058..2175
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     primer_bind     complement(2261..2284)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2611..2630
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      2711..2720
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2720..2743
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2756..2782
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    2798..2822
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(4140..4164)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    4177..4744
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    4194..4746
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4338..4355)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4565..4584
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4758..5357
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4758..4777)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5254..5274
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    5374..5962
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5427..5447)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5845..5856)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             6037..6270
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(6389..6407)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6475..6579
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6580..6648,6649..7440)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(6798..6817)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7611..8199
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8100..8119
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8353..8370
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8610..8631
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8646..8651,8652..8669,8670..8676)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8684..8700
                     /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     8689..8711
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8708..8724
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8708..8724
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     8774..8793
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccgaa ttcacaaatg
     2041 gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa
     2101 agaatagtag acataatagc aacagacata caaactaaag aattacaaaa acaaattaca
     2161 aaaattcaaa attttcgggt ttattacagg gacagcagag atccagtttg gactagcatt
     2221 ctaccgggta ggggaggcgc ttttcccaag gcagtctgga gcatgcgctt tagcagcccc
     2281 gctgggcact tggcgctaca caagtggcct ctggcctcgc acacattcca catccaccgg
     2341 taggcgccaa ccggctccgt tctttggtgg ccccttcgcg ccaccttcta ctcctcccct
     2401 agtcaggaag ttcccccccg ccccgcagct cgcgtcgtgc aggacgtgac aaatggaagt
     2461 agcacgtctc actagtctcg tgcagatgga cagcaccgct gagcaatgga agcgggtagg
     2521 cctttggggc agcggccaat agcagctttg ctccttcgct ttctgggctc aggggcgggg
     2581 cgggcgcccg aaggtcctcc ggaggcccgg cattctgcac gcttcaaaag cgcacgtctg
     2641 ccgcgctgtt ctcctcttcc tcatctccgg gcctttcgac ctgcggccgc agatctctcg
     2701 aggttgatct gccaccatgg actacaagga tgacgatgac aagctcgatg gaggataccc
     2761 atacgatgtt ccagattacg ctgctcgagg ttatcaaaca agtttgtaca aaaaagcagg
     2821 cttcaccatg agctcccctg gcaccgagag cgcgggaaag agcctgcagt accgagtgga
     2881 ccacctgctg agcgccgtgg agaatgagct gcaggcgggc agcgagaagg gcgaccccac
     2941 agagcgcgaa ctgcgcgtgg gcctggagga gagcgagctg tggctgcgct tcaaggagct
     3001 caccaatgag atgatcgtga ccaagaacgg caggaggatg tttccggtgc tgaaggtgaa
     3061 cgtgtctggc ctggacccca acgccatgta ctccttcctg ctggacttcg tggcggcgga
     3121 caaccaccgc tggaagtacg tgaacgggga atgggtgccg gggggcaagc cggagccgca
     3181 ggcgcccagc tgcgtctaca tccaccccga ctcgcccaac ttcggggccc actggatgaa
     3241 ggctcccgtc tccttcagca aagtcaagct caccaacaag ctcaacggag ggggccagat
     3301 catgctgaac tccttgcata agtatgagcc tcgaatccac atagtgagag ttgggggtcc
     3361 acagcgcatg atcaccagcc actgcttccc tgagacccag ttcatagctg tgactgctta
     3421 tcagaacgag gagatcacag ctcttaaaat taagtacaat ccatttgcaa aagctttcct
     3481 tgatgcaaag gaaagaagtg atcacaaaga gatgatggag gaacccggag acagccagca
     3541 acctgggtac tcccaatcag gggggtggct tcttcctgga accagcaccc tgtgtccacc
     3601 tgcaaatcct catcctcagt ttggaggtgc cctctccctc ccctccacgc acagctgtga
     3661 caggtaccca accctgagga gccaccggtc ctcaccctac cccagcccct atgctcatcg
     3721 gaacaattct ccaacctatt ctgacaactc acctgcatgt ttatccatgc tgcaatccca
     3781 tgacaattgg tccagccttg gaatgcctgc ccatcccagc atgctccccg tgagccacaa
     3841 tgccagccca cctaccagct ccagtcagta ccccagcctg tggtctgtga gcaacggcgc
     3901 cgtcaccccg ggctcccagg cagcagccgt gtccaacggg ctgggggccc agttcttccg
     3961 gggctccccc gcgcactaca cacccctcac ccatccggtc tcggcgccct cttcctcggg
     4021 atccccactg tacgaagggg cggccgcggc cacagacatc gtggacagcc agtacgacgc
     4081 cgcagcccaa ggccgcctca tagcctcatg gacacctgtg tcgccacctt ccatgtgaga
     4141 cccagctttc ttgtacaaag tggttcgata acgaattccg cccccccccc ctaacgttac
     4201 tggccgaagc cgcttggaat aaggccggtg tgcgtttgtc tatatgttat tttccaccat
     4261 attgccgtct tttggcaatg tgagggcccg gaaacctggc cctgtcttct tgacgagcat
     4321 tcctaggggt ctttcccctc tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga
     4381 agcagttcct ctggaagctt cttgaagaca aacaacgtct gtagcgaccc tttgcaggca
     4441 gcggaacccc ccacctggcg acaggtgcct ctgcggccaa aagccacgtg tataagatac
     4501 acctgcaaag gcggcacaac cccagtgcca cgttgtgagt tggatagttg tggaaagagt
     4561 caaatggctc tcctcaagcg tattcaacaa ggggctgaag gatgcccaga aggtacccca
     4621 ttgtatggga tctgatctgg ggcctcggtg cacatgcttt acatgtgttt agtcgaggtt
     4681 aaaaaaacgt ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg
     4741 ataatatggc cacaaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg
     4801 acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc
     4861 acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca
     4921 cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg
     4981 tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca
     5041 tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc
     5101 cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc
     5161 agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg
     5221 gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg
     5281 gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc
     5341 gcaagcccgg tgcctgatgc atcgatggat cctaatcaac ctctggatta caaaatttgt
     5401 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     5461 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     5521 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     5581 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     5641 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     5701 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     5761 tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc
     5821 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     5881 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     5941 tccctttggg ccgcctcccc gcctgagatc ctttaagacc aatgacttac aaggcagctg
     6001 tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt cactcccaac
     6061 gaagacaaga tctgcttttt gcttgtactg ggtctctctg gttagaccag atctgagcct
     6121 gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag
     6181 tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac
     6241 ccttttagtc agtgtggaaa atctctagca gtagtagttc atgtcatctt attattcagt
     6301 atttataact tgcaaagaaa tgaatatcag agagtgagag gcccgggtta attaaggaaa
     6361 gggctagatc attcttgaag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     6421 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     6481 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     6541 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     6601 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     6661 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     6721 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     6781 agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag
     6841 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     6901 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     6961 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     7021 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     7081 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     7141 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     7201 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     7261 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     7321 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     7381 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     7441 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     7501 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     7561 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     7621 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     7681 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     7741 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     7801 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     7861 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     7921 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     7981 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     8041 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     8101 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     8161 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     8221 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     8281 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     8341 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg
     8401 cctctccccg cgcgttggcc gattcattaa tgcagcaagc tcatggctga ctaatttttt
     8461 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     8521 gcttttttgg aggcctaggc ttttgcaaaa agctccccgt ggcacgacag gtttcccgac
     8581 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     8641 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     8701 tttcacacag gaaacagcta tgacatgatt acgaatttca caaataaagc atttttttca
     8761 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatcaac
     8821 tggataactc aagctaacca aaatcatccc aaacttccca ccccataccc tattaccact
     8881 gccaattacc tgtggtttca tttactctaa acctgtgatt cctctgaatt attttcattt
     8941 taaagaaatt gtatttgtta aatatgtact acaaacttag tagt
//
