LOCUS       sequence-388976-        8282 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral expression of candidate orphan TCR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8282)
  AUTHORS   Akama-Garren EH, Yin X, Prestwood TR, Ma M, Utz PJ, Carroll MC
  TITLE     T cell help shapes B cell tolerance.
  JOURNAL   Sci Immunol. 2024 Feb 16;9(92):eadj7029. doi: 
            10.1126/sciimmunol.adj7029. Epub 2024 Feb 16.
  PUBMED    38363829
REFERENCE   2  (bases 1 to 8282)
  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: "Sci 
            Immunol."; date: "2024-02-16"; volume: "9(92):eadj7029. doi"; pages:
            " 10.1126/sciimmunol.adj7029. Epub 2024 Feb 16"
COMMENT     Sequence Label: pMIGII-TCR-C
FEATURES             Location/Qualifiers
     source          1..8282
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     5..27
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     155..174
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             374..890
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    954..1295
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             1362..1778
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1708..1730
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1746..1762
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     CDS             2602..2658
                     /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    3617..4166
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3761..3778)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3988..4007
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(4169..4171,4172..4174,4175..4888)
                     /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(4169..4171,4172..4174,4175..4885)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4214..4235)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4475..4494)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4822..4843
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             5050..5564
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5726..5742
                     /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(5750..5756,5757..5774,5775..5780))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5795..5816
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5933..5950)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6104..6692)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6184..6203)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6863..7654,7655..7723))
                     /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     7486..7505
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7724..7828)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7896..7914
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7952..7974)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     8074..8093
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ttttcccagt cacgacgttg taaaacgacg gcgcaaggaa tggtgcatgc aaggagatgg
       61 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca
      121 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag
      181 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggcgattag
      241 tccaatttgt taaagacagg atatcagtgg tccaggctct agttttgact caacaatatc
      301 accagctgaa gcctatagag tacgagccat agataaaata aaagatttta tttagtctcc
      361 agaaaaaggg gggaatgaaa gaccccacct gtaggtttgg caagctagct taagtaacgc
      421 cattttgcaa ggcatggaaa atacataact gagaatagag aagttcagat caaggttagg
      481 aacagagaga cagcagaata tgggccaaac aggatatctg tggtaagcag ttcctgcccc
      541 ggctcagggc caagaacaga tggtccccag atgcggtccc gccctcagca gtttctagag
      601 aaccatcaga tgtttccagg gtgccccaag gacctgaaat gaccctgtgc cttatttgaa
      661 ctaaccaatc agttcgcttc tcgcttctgt tcgcgcgctt ctgctccccg agctcaataa
      721 aagagcccac aacccctcac tcggcgcgcc agtcctccga tagactgcgt cgcccgggta
      781 cccgtattcc caataaagcc tcttgctgtt tgcatccgaa tcgtggactc gctgatcctt
      841 gggagggtct cctcagattg attgactgcc cacctcgggg gtctttcatt tggaggttcc
      901 accgagattt ggagacccct gcccagggac caccgacccc cccgccggga ggtaagctgg
      961 ccagcggtcg tttcgtgtct gtctctgtct ttgtgcgtgt ttgtgccggc atctaatgtt
     1021 tgcgcctgcg tctgtactag ttagctaact agctctgtat ctggcggacc cgtggtggaa
     1081 ctgacgagtt ctgaacaccc ggccgcaacc ctgggagacg tcccagggac tttgggggcc
     1141 gtttttgtgg cccgacctga ggaagggagt cgatgtggaa tccgaccccg tcaggatatg
     1201 tggttctggt aggagacgag aacctaaaac agttcccgcc tccgtctgaa tttttgcttt
     1261 cggtttggaa ccgaagccgc gcgtcttgtc tgctgcagcg ctgcagcatc gttctgtgtt
     1321 gtctctgtct gactgtgttt ctgtatttgt ctgaaaatta gggccagact gttaccactc
     1381 ccttaagttt gaccttaggt cactggaaag atgtcgagcg gatcgctcac aaccagtcgg
     1441 tagatgtcaa gaagagacgt tgggttacct tctgctctgc agaatggcca acctttaacg
     1501 tcggatggcc gcgagacggc acctttaacc gagacctcat cacccaggtt aagatcaagg
     1561 tcttttcacc tggcccgcat ggacacccag accaggtccc ctacatcgtg acctgggaag
     1621 ccttggcttt tgacccccct ccctgggtca agccctttgt acaccctaag cctccgcctc
     1681 ctcttcctcc atccgccccg tctctccccc ttgaacctcc tcgttcgacc ccgcctcgat
     1741 cctcccttta tccagccctc actccttctc taggcgccgg aattcagatc taccatgcag
     1801 aggaacctgg gagctgtgct ggggattctg tgggtgcaga tttgctgggt gagaggagat
     1861 caggtggagc agagtccttc agccctgagc ctccacgagg gaaccggttc tgctctgaga
     1921 tgcaatttta ctaccaccat gagggctgtg cagtggttcc gaaagaattc caggggcagc
     1981 ctcatcaatc tgttctactt ggcttcagga acaaaggaga atgggaggct aaagtcagca
     2041 tttgattcta aggagcgcta cagcaccctg cacatcaggg atgcccagct ggaggactca
     2101 ggcacttact tctgtgctgc tggtgcatct tctggcagct ggcaactcat ctttggatct
     2161 ggaacccaac tgacagttat gcctgacatc cagaacccag aacctgctgt gtaccagtta
     2221 aaagatcctc ggtctcagga cagcaccctc tgcctgttca ccgactttga ctcccaaatc
     2281 aatgtgccga aaaccatgga atctggaacg ttcatcactg acaaaactgt gctggacatg
     2341 aaagctatgg attccaagag caatggggcc attgcctgga gcaaccagac aagcttcacc
     2401 tgccaagata tcttcaaaga gaccaacgcc acctacccca gttcagacgt tccctgtgat
     2461 gccacgttga ctgagaaaag ctttgaaaca gatatgaacc taaactttca aaacctgtca
     2521 gttatgggac tccgaatcct cctgctgaaa gtagccggat ttaacctgct catgacgctg
     2581 aggctgtggt ccggctccgg agccacgaac ttctctctgt taaagcaagc aggagacgtg
     2641 gaagaaaacc ccggtcccat gggctccagg ctctttctgg tcttgagcct cctgtgtaca
     2701 aaacacatgg aggctgcagt cacccaaagc cctagaaaca aggtgacagt aacaggagga
     2761 aacgtgacat tgagctgtcg ccagactaat agccacaact acatgtactg gtatcggcag
     2821 gacactgggc atgggctgag gctgatccat tactcatatg gtgctggcaa ccttcaaata
     2881 ggagatgtcc ctgatgggta caaggccacc agaacaacgc aagaagactt cttcctcctg
     2941 ctggaattgg cttctccctc tcagacatct ttgtacttct gtgccagcag tgagggactg
     3001 gggagctcct atgaacagta cttcggtccc ggcaccaggc tcacggtttt agaggatctg
     3061 agaaatgtga ctccacccaa ggtctccttg tttgagccat caaaagcaga gattgcaaac
     3121 aaacaaaagg ctaccctcgt gtgcttggcc aggggcttct tccctgacca cgtggagctg
     3181 agctggtggg tgaatggcaa ggaggtccac agtggggtca gcacggaccc tcaggcctac
     3241 aaggagagca attatagcta ctgcctgagc agccgcctga gggtctctgc taccttctgg
     3301 cacaatcctc gaaaccactt ccgctgccaa gtgcagttcc atgggctttc agaggaggac
     3361 aagtggccag agggctcacc caaacctgtc acacagaaca tcagtgcaga ggcctggggc
     3421 cgagcagact gtggaatcac ttcagcatcc tatcatcagg gggttctgtc tgcaaccatc
     3481 ctctatgaga tcctactggg gaaggccacc ctatatgctg tgctggtcag tggcctggtg
     3541 ctgatggcca tggtcaagaa aaaaaattcc tgactcgagc aattgctcga gcgggatcaa
     3601 ttccgccccc cccctaacgt tactggccga agccgcttgg aataaggccg gtgtgcgttt
     3661 gtctatatgt tattttccac catattgccg tcttttggca atgtgagggc ccggaaacct
     3721 ggccctgtct tcttgacgag cattcctagg ggtctttccc ctctcgccaa aggaatgcaa
     3781 ggtctgttga atgtcgtgaa ggaagcagtt cctctggaag cttcttgaag acaaacaacg
     3841 tctgtagcga ccctttgcag gcagcggaac cccccacctg gcgacaggtg cctctgcggc
     3901 caaaagccac gtgtataaga tacacctgca aaggcggcac aaccccagtg ccacgttgtg
     3961 agttggatag ttgtggaaag agtcaaatgg ctctcctcaa gcgtattcaa caaggggctg
     4021 aaggatgccc agaaggtacc ccattgtatg ggatctgatc tggggcctcg gtgcacatgc
     4081 tttacatgtg tttagtcgag gttaaaaaaa cgtctaggcc ccccgaacca cggggacgtg
     4141 gttttccttt gaaaaacacg ataataccat ggtgagcaag ggcgaggagc tgttcaccgg
     4201 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
     4261 cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
     4321 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg
     4381 cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     4441 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     4501 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     4561 caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt
     4621 ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa
     4681 catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga
     4741 cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga
     4801 ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac
     4861 tctcggcatg gacgagctgt acaagtaaag cggccgcgac tctagagtcg acctgcaggc
     4921 atgcaagctt caggtagccg gctaacgtta acaaccggta cctctagaac tatagctagc
     4981 atgcgcaaat ttaaagcgct gatatcgata aaataaaaga ttttatttag tctccagaaa
     5041 aaggggggaa tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt
     5101 tgcaaggcat ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag
     5161 agagacagca gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc
     5221 agggccaaga acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca
     5281 tcagatgttt ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac
     5341 caatcagttc gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag
     5401 cccacaaccc ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt
     5461 gtatccaata aaccctcttg cagttgcatc cgacttgtgg tctcgctgtt ccttgggagg
     5521 gtctcctctg agtgattgac tacccgtcag cgggggtctt tcatgggtaa cagtttcttg
     5581 aagttggaga acaacattct gagggtagga gtcgaatatt aagtaatcct gactcaatta
     5641 gccactgttt tgaatccaca tactccaata ctcctgaaat agttcattat ggacagcgca
     5701 gaaagagctg gggagaattg tgaaattgtt atccgctcac aattccacac aacatacgag
     5761 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     5821 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     5881 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     5941 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     6001 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     6061 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     6121 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     6181 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     6241 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     6301 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     6361 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     6421 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     6481 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     6541 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     6601 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     6661 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     6721 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     6781 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     6841 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     6901 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     6961 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     7021 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     7081 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     7141 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     7201 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     7261 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     7321 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     7381 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     7441 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     7501 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     7561 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     7621 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     7681 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     7741 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     7801 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct
     7861 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     7921 gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg
     7981 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg
     8041 gtgttggcgg gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag
     8101 tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc
     8161 gccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc
     8221 tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag
     8281 gg
//
