LOCUS       sequence-382622-        8261 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 8261)
  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 8261)
  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-F
FEATURES             Location/Qualifiers
     source          1..8261
                     /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             2593..2649
                     /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    3596..4145
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3740..3757)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3967..3986
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(4148..4150,4151..4153,4154..4867)
                     /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(4148..4150,4151..4153,4154..4864)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4193..4214)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4454..4473)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4801..4822
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             5029..5543
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5705..5721
                     /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(5729..5735,5736..5753,5754..5759))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5774..5795
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5912..5929)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6083..6671)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6163..6182)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6842..7633,7634..7702))
                     /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     7465..7484
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7703..7807)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7875..7893
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7931..7953)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     8053..8072
                     /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 cgaccaccat gagggctgtg cagtggttcc aacagaattc caggggcagc
     1981 ctcatcaatc tgttctactt ggcttcagga acaaaggaga atgggaggtt aaagtcaaca
     2041 ttcaattcta aggagagcta cagcaccctg cacatcaggg atgcccagct ggaggactca
     2101 ggcacttact tctgtgctgc tgaatttgca aacaagatga tctttggctt gggaaccatt
     2161 ttgagagtca gacctcacat ccagaaccca gaacctgctg tgtaccagtt aaaagatcct
     2221 cggtctcagg acagcaccct ctgcctgttc accgactttg actcccaaat caatgtgccg
     2281 aaaaccatgg aatctggaac gttcatcact gacaaaactg tgctggacat gaaagctatg
     2341 gattccaaga gcaatggggc cattgcctgg agcaaccaga caagcttcac ctgccaagat
     2401 atcttcaaag agaccaacgc cacctacccc agttcagacg ttccctgtga tgccacgttg
     2461 actgagaaaa gctttgaaac agatatgaac ctaaactttc aaaacctgtc agttatggga
     2521 ctccgaatcc tcctgctgaa agtagccgga tttaacctgc tcatgacgct gaggctgtgg
     2581 tccggctccg gagccacgaa cttctctctg ttaaagcaag caggagacgt ggaagaaaac
     2641 cccggtccca tgggcatcca gaccctctgt tgtgtgatct tttatgttct gatagcaaat
     2701 cacacagatg ctggagttac ccagacaccc agacatgagg tggcagagaa aggacaaaca
     2761 ataatcctga agtgtgagcc agtttcaggc cacaatgacc ttttctggta cagacagacc
     2821 aagatacagg gactagagtt gctgagctac ttccgcagca agtctcttat ggaagatggt
     2881 ggggctttca aggatcgatt caaagctgag atgctaaatt catccttctc cactctgaag
     2941 attcaaccta cagaacccaa ggactcagct gtgtatctgt gtgccagcag tttagggggg
     3001 tatgaacagt acttcggtcc cggcaccagg ctcacggttt tagaggatct gagaaatgtg
     3061 actccaccca aggtctcctt gtttgagcca tcaaaagcag agattgcaaa caaacaaaag
     3121 gctaccctcg tgtgcttggc caggggcttc ttccctgacc acgtggagct gagctggtgg
     3181 gtgaatggca aggaggtcca cagtggggtc agcacggacc ctcaggccta caaggagagc
     3241 aattatagct actgcctgag cagccgcctg agggtctctg ctaccttctg gcacaatcct
     3301 cgaaaccact tccgctgcca agtgcagttc catgggcttt cggaggagga caagtggcca
     3361 gagggctcac ccaaacctgt cacacagaac atcagtgcag aggcctgggg ccgagcagac
     3421 tgtggaatca cttcagcatc ctatcatcag ggggttctgt ctgcaaccat cctctatgag
     3481 atcctactgg ggaaggccac cctatatgct gtgctggtca gtggcctggt gctgatggcc
     3541 atggtcaaga gaaaaaattc ctgactcgag cgggatcaat tccgcccccc ccctaacgtt
     3601 actggccgaa gccgcttgga ataaggccgg tgtgcgtttg tctatatgtt attttccacc
     3661 atattgccgt cttttggcaa tgtgagggcc cggaaacctg gccctgtctt cttgacgagc
     3721 attcctaggg gtctttcccc tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag
     3781 gaagcagttc ctctggaagc ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg
     3841 cagcggaacc ccccacctgg cgacaggtgc ctctgcggcc aaaagccacg tgtataagat
     3901 acacctgcaa aggcggcaca accccagtgc cacgttgtga gttggatagt tgtggaaaga
     3961 gtcaaatggc tctcctcaag cgtattcaac aaggggctga aggatgccca gaaggtaccc
     4021 cattgtatgg gatctgatct ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg
     4081 ttaaaaaaac gtctaggccc cccgaaccac ggggacgtgg ttttcctttg aaaaacacga
     4141 taataccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     4201 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     4261 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     4321 gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca
     4381 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     4441 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     4501 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     4561 ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca
     4621 gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     4681 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     4741 caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     4801 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     4861 caagtaaagc ggccgcgact ctagagtcga cctgcaggca tgcaagcttc aggtagccgg
     4921 ctaacgttaa caaccggtac ctctagaact atagctagca tgcgcaaatt taaagcgctg
     4981 atatcgataa aataaaagat tttatttagt ctccagaaaa aggggggaat gaaagacccc
     5041 acctgtaggt ttggcaagct agcttaagta acgccatttt gcaaggcatg gaaaatacat
     5101 aactgagaat agagaagttc agatcaaggt taggaacaga gagacagcag aatatgggcc
     5161 aaacaggata tctgtggtaa gcagttcctg ccccggctca gggccaagaa cagatggtcc
     5221 ccagatgcgg tcccgccctc agcagtttct agagaaccat cagatgtttc cagggtgccc
     5281 caaggacctg aaatgaccct gtgccttatt tgaactaacc aatcagttcg cttctcgctt
     5341 ctgttcgcgc gcttctgctc cccgagctca ataaaagagc ccacaacccc tcactcggcg
     5401 cgccagtcct ccgatagact gcgtcgcccg ggtacccgtg tatccaataa accctcttgc
     5461 agttgcatcc gacttgtggt ctcgctgttc cttgggaggg tctcctctga gtgattgact
     5521 acccgtcagc gggggtcttt catgggtaac agtttcttga agttggagaa caacattctg
     5581 agggtaggag tcgaatatta agtaatcctg actcaattag ccactgtttt gaatccacat
     5641 actccaatac tcctgaaata gttcattatg gacagcgcag aaagagctgg ggagaattgt
     5701 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     5761 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     5821 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     5881 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     5941 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     6001 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     6061 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     6121 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     6181 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     6241 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     6301 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     6361 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     6421 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     6481 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     6541 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     6601 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     6661 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     6721 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     6781 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     6841 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     6901 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     6961 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     7021 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     7081 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     7141 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     7201 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     7261 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     7321 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     7381 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     7441 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     7501 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     7561 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     7621 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     7681 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     7741 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     7801 ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga
     7861 cattaaccta taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg
     7921 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
     7981 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct
     8041 ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa
     8101 taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc
     8161 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
     8221 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg g
//
