LOCUS       sequence-228424-        9478 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  stable TCR expression in human T cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9478)
  AUTHORS   Wang R, Pscheid R, Ghumra A, Kan LYL, Cochrane S, Fairclough L, 
            Alcocer MJC
  TITLE     Towards a surrogate system to express human lipid binding TCRs.
  JOURNAL   Biotechnol Lett. 2019 Oct;41(10):1095-1104. doi: 
            10.1007/s10529-019-02713-2. Epub 2019 Jul 26.
  PUBMED    31346817
REFERENCE   2  (bases 1 to 9478)
  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"; doi: 
            "10.1007/s10529-019-02713-2"; journalName: "Biotechnol Lett"; date: 
            "2019-10"; volume: "41"; issue: "10"; pages: "1095-1104"
COMMENT     Sequence Label: pMJA289= pSin-EF2-TRAV10-P2A-TRBV25-Puro
FEATURES             Location/Qualifiers
     source          1..9478
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             117..750
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    797..922
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1415..1648
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1833..1877
                     /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    2164..2281
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2410..3588
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2641..3579
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3536..3556
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             4493..4549
                     /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"
     misc_feature    join(5594..6168,6169..6171,6172..6180)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(5761..5778)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     5988..6007
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             6200..6799
                     /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(6200..6219)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6696..6716
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             7002..7235
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(7386..7403)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7392..7616
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     regulatory      7659..7668
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        7712..7816
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7817..7885,7886..8677)
                     /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(8035..8054)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8848..9436
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9337..9356
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cggaccgcca ctgccaatta cctgtggttt catttactct aaacctgtga ttcctctgaa
       61 ttattttcat tttaaagaaa ttgtatttgt taaatatgta ctacaaactt agtagttgga
      121 agggctaatt cactcccaaa gaagacaaga tatccttgat ctgtggatct accacacaca
      181 aggctacttc cctgattagc agaactacac accagggcca ggggtcagat atccactgac
      241 ctttggatgg tgctacaagc tagtaccagt tgagccagat aaggtagaag aggccaataa
      301 aggagagaac accagcttgt tacaccctgt gagcctgcat gggatggatg acccggagag
      361 agaagtgtta gagtggaggt ttgacagccg cctagcattt catcacgtgg cccgagagct
      421 gcatccggag tacttcaaga actgctgata tcgagcttgc tacaagggac tttccgctgg
      481 ggactttcca gggaggcgtg gcctgggcgg gactggggag tggcgagccc tcagatcctg
      541 catataagca gctgcttttt gcctgtactg ggtctctctg gttagaccag atctgagcct
      601 gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag
      661 tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac
      721 ccttttagtc agtgtggaaa atctctagca gtggcgcccg aacagggact tgaaagcgaa
      781 agggaaacca gaggagctct ctcgacgcag gactcggctt gctgaagcgc gcacggcaag
      841 aggcgagggg cggcgactgg tgagtacgcc aaaaattttg actagcggag gctagaagga
      901 gagagatggg tgcgagagcg tcagtattaa gcgggggaga attagatcgc gatgggaaaa
      961 aattcggtta aggccagggg gaaagaaaaa atataaatta aaacatatag tatgggcaag
     1021 cagggagcta gaacgattcg cagttaatcc tggcctgtta gaaacatcag aaggctgtag
     1081 acaaatactg ggacagctac aaccatccct tcagacagga tcagaagaac ttagatcatt
     1141 atataataca gtagcaaccc tctattgtgt gcatcaaagg atagagataa aagacaccaa
     1201 ggaagcttta gacaagatag aggaagagca aaacaaaagt aagaccaccg cacagcaagc
     1261 ggccgctgat cttcagacct ggaggaggag atatgaggga caattggaga agtgaattat
     1321 ataaatataa agtagtaaaa attgaaccat taggagtagc acccaccaag gcaaagagaa
     1381 gagtggtgca gagagaaaaa agagcagtgg gaataggagc tttgttcctt gggttcttgg
     1441 gagcagcagg aagcactatg ggcgcagcgt caatgacgct gacggtacag gccagacaat
     1501 tattgtctgg tatagtgcag cagcagaaca atttgctgag ggctattgag gcgcaacagc
     1561 atctgttgca actcacagtc tggggcatca agcagctcca ggcaagaatc ctggctgtgg
     1621 aaagatacct aaaggatcaa cagctcctgg ggatttgggg ttgctctgga aaactcattt
     1681 gcaccactgc tgtgccttgg aatgctagtt ggagtaataa atctctggaa cagatttgga
     1741 atcacacgac ctggatggag tgggacagag aaattaacaa ttacacaagc ttaatacact
     1801 ccttaattga agaatcgcaa aaccagcaag aaaagaatga acaagaatta ttggaattag
     1861 ataaatgggc aagtttgtgg aattggttta acataacaaa ttggctgtgg tatataaaat
     1921 tattcataat gatagtagga ggcttggtag gtttaagaat agtttttgct gtactttcta
     1981 tagtgaatag agttaggcag ggatattcac cattatcgtt tcagacccac ctcccaaccc
     2041 cgaggggacc cgacaggccc gaaggaatag aagaagaagg tggagagaga gacagagaca
     2101 gatccattcg attagtgaac ggatctcgac ggtatcgcca caaatggcag tattcatcca
     2161 caattttaaa agaaaggggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
     2221 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
     2281 tcgggtttat tacagggaca gcagagatcc actttggatc gataagcttt gcaaagatgg
     2341 ataaagtttt aaacagagag gaatctttgc agctaatgga ccttctaggt cttgaaagga
     2401 gtgggaattg gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag
     2461 aagttggggg gaggggtcgg caattgaacc ggtgcctaga gaaggtggcg cggggtaaac
     2521 tgggaaagtg atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat
     2581 ataagtgcag tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag
     2641 gtaagtgccg tgtgtggttc ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg
     2701 ccttgaatta cttccacctg gctgcagtac gtgattcttg atcccgagct tcgggttgga
     2761 agtgggtggg agagttcgag gccttgcgct taaggagccc cttcgcctcg tgcttgagtt
     2821 gaggcctggc ctgggcgctg gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt
     2881 ctcgctgctt tcgataagtc tctagccatt taaaattttt gatgacctgc tgcgacgctt
     2941 tttttctggc aagatagtct tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt
     3001 tttggggccg cgggcggcga cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg
     3061 ggcctgcgag cgcggccacc gagaatcgga cgggggtagt ctcaagctgg ccggcctgct
     3121 ctggtgcctg gcctcgcgcc gccgtgtatc gccccgccct gggcggcaag gctggcccgg
     3181 tcggcaccag ttgcgtgagc ggaaagatgg ccgcttcccg gccctgctgc agggagctca
     3241 aaatggagga cgcggcgctc gggagagcgg gcgggtgagt cacccacaca aaggaaaagg
     3301 gcctttccgt cctcagccgt cgcttcatgt gactccacgg agtaccgggc gccgtccagg
     3361 cacctcgatt agttctcgag cttttggagt acgtcgtctt taggttgggg ggaggggttt
     3421 tatgcgatgg agtttcccca cactgagtgg gtggagactg aagttaggcc agcttggcac
     3481 ttgatgtaat tctccttgga atttgccctt tttgagtttg gatcttggtt cattctcaag
     3541 cctcagacag tggttcaaag tttttttctt ccatttcagg tgtcgtgagg aattgatcct
     3601 tcgaactagt atgaaacaag gcagcggatt tgaggcaggc ttctccagaa caaccatgaa
     3661 gaagcacctg accacctttc tcgtgatcct gtggctgtac ttctacagag gcaacggcaa
     3721 gaaccaggtg gaacagagcc cccagagcct gatcatcctg gaaggcaaga actgcaccct
     3781 gcagtgcaac tacaccgtgt ccccattcag caacctgaga tggtacaagc aggacaccgg
     3841 cagaggcccc gtgtccctga ccatcatgac cttcagcgag aacaccaaga gcaacggcag
     3901 atacaccgcc accctggacg ccgacacaaa gcagagcagc ctgcacatca ccgccagcca
     3961 gctgagcgac agcgccagct acatctgcgt ggtgtccgac agaggcagca ccctgggcag
     4021 actgtacttc ggcagaggaa cccagctgac cgtgtggccc aacatccaga accctgaccc
     4081 tgccgcgtac cagctgagag actccaagtc cagcgacaag agcgtgtgcc tgttcaccga
     4141 cttcgacagc cagaccaacg tgtcccagag caaggacagc gacgtgtaca tcaccgacaa
     4201 gaccgtgctg gacatgagat ccatggactt caagtctaac agcgccgtgg cctggtccaa
     4261 caagagcgac ttcgcctgcg ccaacgcctt caacaacagc attatccccg aggacacatt
     4321 cttcccaagc cccgagagca gctgcgacgt gaagctggtg gaaaagagct tcgagacaga
     4381 caccaacctg aacttccaga acctgagcgt gatcggcttc agaatcctgc tgctgaaggt
     4441 ggccggcttc aacctgctga tgaccctgag actgtggtcc agcggaagcg gagctactaa
     4501 ctttagcctg ctgaagcagg ctggagatgt ggaggagaac cctggaccta tgaccatcag
     4561 actgctgtgc tacgtgggct tctacttcct gggcgctggc ctgatggaag ccgacatcta
     4621 ccagaccccc agatacctcg tgatcggcac cggcaagaaa atcaccctgg aatgcagcca
     4681 gaccatgggc cacgacaaga tgtactggta tcagcaggac cccggcatgg aactgcacct
     4741 gatccactac agctacggcg tgaacagcac cgagaagggc gacctgagca gcgagagcac
     4801 cgtgtccaga atcagaaccg agcacttccc actgaccctg gaaagcgcta gacccagcca
     4861 caccagccag tacctgtgtg ccagcagctc tcctctgggc ggaacacctg gcgccaacgt
     4921 gctgacattt ggcgctggca gcagactgac cgtgctggaa gatctgaaga acgtgttccc
     4981 acccgaggtg gccgtgttcg agccttctga ggccgagatc tcccacaccc agaaagccac
     5041 cctcgtgtgt ctggccaccg gcttctaccc cgaccacgtg gaactgtctt ggtgggtcaa
     5101 cggcaaagag gtgcacagcg gcgtgtccac cgatccccag cctctgaaag agcagcccgc
     5161 cctgaacgac agcagatact gcctgtccag caggctgaga gtgtccgcca ccttctggca
     5221 gaaccccaga aaccacttca gatgccaggt gcagttttac ggcctgagcg agaacgacga
     5281 gtggacccag gacagagcca agcccgtgac ccagatcgtg tctgctgagg cctggggcag
     5341 agccgactgt ggcttcacaa gcgagagcta ccagcagggc gtgctgagcg ccaccatcct
     5401 gtacgagatc ctgctgggca aggccaccct gtacgccgtg ctggtgtctg ccctggtgct
     5461 gatggccatg gtcaagagaa aggacagcag aggctgagaa ttcgaagctt gagctcgaga
     5521 tctgagtccg gtagcgctag ttcgaaggat ccgcatgcat ctagggcggc caattccgcc
     5581 cctctccccc ccccccctct ccctcccccc cccctaacgt tactggccga agccgcttgg
     5641 aataaggccg gtgtgcgttt gtctatatgt tattttccac catattgccg tcttttggca
     5701 atgtgagggc ccggaaacct ggccctgtct tcttgacgag cattcctagg ggtctttccc
     5761 ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa ggaagcagtt cctctggaag
     5821 cttcttgaag acaaacaacg tctgtagcga ccctttgcag gcagcggaac cccccacctg
     5881 gcgacaggtg cctctgcggc caaaagccac gtgtataaga tacacctgca aaggcggcac
     5941 aaccccagtg ccacgttgtg agttggatag ttgtggaaag agtcaaatgg ctctcctcaa
     6001 gcgtattcaa caaggggctg aaggatgccc agaaggtacc ccattgtatg ggatctgatc
     6061 tggggcctcg gtgcacatgc tttacatgtg tttagtcgag gttaaaaaaa cgtctaggcc
     6121 ccccgaacca cggggacgtg gttttccttt gaaaaacacg atgataagct tgccacaacc
     6181 cacaaggaga cgaccttcca tgaccgagta caagcccacg gtgcgcctcg ccacccgcga
     6241 cgacgtcccc cgggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg
     6301 ccacaccgtc gacccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct
     6361 cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc
     6421 ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg
     6481 catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc
     6541 gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca
     6601 ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc
     6661 cggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct acgagcggct
     6721 cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac
     6781 ccgcaagccc ggtgcctgat aataggcggc cgctcgagac ctagaaaaac atggagcaat
     6841 cacaagtagc aatacagcag ctaccaatgc tgattgtgcc tggctagaag cacaagagga
     6901 ggaggaggtg ggttttccag tcacacctca ggtaccttta agaccaatga cttacaaggc
     6961 agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc
     7021 ccaacgaaga caagatctgc tttttgcttg tactgggtct ctctggttag accagatctg
     7081 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
     7141 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
     7201 cagacccttt tagtcagtgt ggaaaatctc tagcagtagt agttcatgtc atcttattat
     7261 tcagtattta taacttgcaa agaaatgaat atcagagagt gagaggcctt gacattataa
     7321 tagatttagc aggaattgaa ctaggagtgg agcacacagg caaagttcta gagctcgctg
     7381 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     7441 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     7501 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     7561 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     7621 tgaggcggaa agaaccagct gggggcgcgc ccctcgaggc cgccatggtc atagctgttt
     7681 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     7741 aatacattca aatatgtatc cgctcacgag acaataaccc tgataaatgc ttcaataata
     7801 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     7861 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     7921 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     7981 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     8041 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     8101 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     8161 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     8221 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     8281 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     8341 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     8401 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     8461 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     8521 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     8581 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     8641 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     8701 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     8761 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     8821 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     8881 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     8941 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     9001 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     9061 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     9121 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     9181 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     9241 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     9301 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     9361 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     9421 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttctggc cttttgca
//
