LOCUS       sequence-482724-        7438 bp    DNA     circular SYN 08-AUG-2025
DEFINITION  Destination vector for complete TCR library.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7438)
  AUTHORS   Gaglione SA, Mukkamala RS, Krishna C, Smith BE, Wadsworth II MH, 
            Jelinsky SA, Perez CR, Schmidt-Hong L, Katz EL, Gellatly KJ, Ali LR,
            Shen J, Holec PV, Zhao QH, Chan AO , Xu EJK, Kravarik KM, Guzova JA,
            Dobson CS, Singh H, Garber M, Dougan M, Dougan SK, Harris JE, 
            Winkler A, Birnbaum ME
  TITLE     Scalable TCR synthesis and screening enables antigen reactivity 
            mapping in vitiligo
  JOURNAL   bioRxiv 2025.04.24.649726
REFERENCE   2  (bases 1 to 7438)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            2025.04.24.649726"
COMMENT     Sequence Label: pHIV-LacZ-SapI
FEATURES             Location/Qualifiers
     source          1..7438
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1684..1917
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2102..2146
                     /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    2444..2561
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2624..3802
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2855..3793
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3750..3770
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    3846..3867
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3882..3887,3888..3905,3906..3912)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3920..3936
                     /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     3925..3947
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3944..3960
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3944..3960
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             3956..4279
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    3973..4029
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(4030..4047)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4030..4046)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4039..4061)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     misc_feature    4312..4715
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4918..4934)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4919..4935)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5444..5624
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(5686..6274)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5766..5785)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6445..7236,7237..7305))
                     /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     7068..7087
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7306..7410)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gaacaaaaca
     1501 aaagtaagac caccgcacag caagcggccg gccgctgatc ttcagacctg gaggaggaga
     1561 tatgagggac aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt
     1621 aggagtagca cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg
     1681 aataggagct ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc
     1741 aatgacgctg acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa
     1801 tttgctgagg gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa
     1861 gcagctccag gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg
     1921 gatttggggt tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg
     1981 gagtaataaa tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga
     2041 aattaacaat tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga
     2101 aaagaatgaa caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa
     2161 cataacaaat tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg
     2221 tttaagaata gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc
     2281 attatcgttt cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga
     2341 agaagaaggt ggagagagag acagagacag atccattcga ttagtgaacg gatcggcact
     2401 gcgtgcgcca attctgcaga caaatggcag tattcatcca caattttaaa agaaaagggg
     2461 ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa
     2521 ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat tacagggaca
     2581 gcagagatcc agtttggtta gtaccgggcc cgctctagcg tgaggctccg gtgcccgtca
     2641 gtgggcagag cgcacatcgc ccacagtccc cgagaagttg gggggagggg tcggcaattg
     2701 aaccggtgcc tagagaaggt ggcgcggggt aaactgggaa agtgatgtcg tgtactggct
     2761 ccgccttttt cccgagggtg ggggagaacc gtatataagt gcagtagtcg ccgtgaacgt
     2821 tctttttcgc aacgggtttg ccgccagaac acaggtaagt gccgtgtgtg gttcccgcgg
     2881 gcctggcctc tttacgggtt atggcccttg cgtgccttga attacttcca cctggctgca
     2941 gtacgtgatt cttgatcccg agcttcgggt tggaagtggg tgggagagtt cgaggccttg
     3001 cgcttaagga gccccttcgc ctcgtgcttg agttgaggcc tggcctgggc gctggggccg
     3061 ccgcgtgcga atctggtggc accttcgcgc ctgtctcgct gctttcgata agtctctagc
     3121 catttaaaat ttttgatgac ctgctgcgac gctttttttc tggcaagata gtcttgtaaa
     3181 tgcgggccaa gatctgcaca ctggtatttc ggtttttggg gccgcgggcg gcgacggggc
     3241 ccgtgcgtcc cagcgcacat gttcggcgag gcggggcctg cgagcgcggc caccgagaat
     3301 cggacggggg tagtctcaag ctggccggcc tgctctggtg cctggcctcg cgccgccgtg
     3361 tatcgccccg ccctgggcgg caaggctggc ccggtcggca ccagttgcgt gagcggaaag
     3421 atggccgctt cccggccctg ctgcagggag ctcaaaatgg aggacgcggc gctcgggaga
     3481 gcgggcgggt gagtcaccca cacaaaggaa aagggccttt ccgtcctcag ccgtcgcttc
     3541 atgtgactcc acggagtacc gggcgccgtc caggcacctc gattagttct cgagcttttg
     3601 gagtacgtcg tctttaggtt ggggggaggg gttttatgcg atggagtttc cccacactga
     3661 gtgggtggag actgaagtta ggccagcttg gcacttgatg taattctcct tggaatttgc
     3721 cctttttgag tttggatctt ggttcattct caagcctcag acagtggttc aaagtttttt
     3781 tcttccattt caggtgtcgt gagcggccgc tgagttaact attctaggca agaagagcca
     3841 ccaattaatg tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg
     3901 gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac
     3961 catgattacg ccaagcttgc atgcctgcag gtcgacccta gaggatcccc gggtaccgag
     4021 ctcaaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca
     4081 acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg
     4141 caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc tgatgcggta
     4201 ttttctcctt acgcatctat gcggtatttc acaccgcata tggtgcactc tcagtacaat
     4261 ctgctctgat gccgcatagt taacgctctt ctatcaattc tgtacaagtg aaatcaacct
     4321 ctggattaca aaatttgtga aagattgact gatattctta actatgttgc tccttttacg
     4381 ctgtgtggat atgctgcttt aatgcctctg tatcatgcta ttgcttcccg tacggctttc
     4441 gttttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt
     4501 gtccgtcaac gtggcgtggt gtgctctgtg tttgctgacg caacccccac tggctggggc
     4561 attgccacca cctgtcaact cctttctggg actttcgctt tccccctccc gatcgccacg
     4621 gcagaactca tcgccgcctg ccttgcccgc tgctggacag gggctaggtt gctgggcact
     4681 gataattccg tggtgttgtc ggggaagctg acgtcctttc catggctgct cgcctgtgtt
     4741 gccaactgga tcctgcgcgg gacgtccttc tgctacgtcc cttcggctct caatccagcg
     4801 gacctccctt cccgaggcct tctgccggtt ctgcggcctc tcccgcgtct tcgctttcgg
     4861 cctccgacga gtcggatctc cctttgggcc gcctccccgc aagtaaagcg gccgcatcga
     4921 taccgtcgac ctcgatcgag acctagaaaa acatggagca atcacaagta gcaatacagc
     4981 agctaccaat gctgattgtg cctggctaga agcacaagag gaggaggagg tgggttttcc
     5041 agtcacacct caggtacctt taagaccaat gacttacaag gcagctgtag atcttagcca
     5101 ctttttaaaa gaaaaggggg gactggaagg gctaattcac tcccaacgaa gacaagatat
     5161 ccttgatctg tggatctacc acacacaagg ctacttccct gattggcaga actacacacc
     5221 agggccaggg atcagatatc cactgacctt tggatggtgc tacaagctag taccagttga
     5281 gcaagagaag gtagaagaag ccaatgaagg agagaacacc cgcttgttac accctgtgag
     5341 cctgcatggg atggatgacc cggagagaga agtattagag tggaggtttg acagccgcct
     5401 agcatttcat cacatggccc gagagctgca tccggactgt actgggtctc tctggttaga
     5461 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata
     5521 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta
     5581 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagcatgt gagcaaaagg
     5641 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     5701 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     5761 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     5821 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     5881 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     5941 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     6001 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     6061 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     6121 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     6181 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     6241 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacgtg
     6301 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     6361 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     6421 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     6481 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     6541 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     6601 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     6661 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     6721 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     6781 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     6841 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     6901 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     6961 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     7021 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     7081 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     7141 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     7201 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     7261 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     7321 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     7381 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgac
//
