LOCUS       sequence-412258-        5632 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  This plasmid can be used to generate HDR templates for non-viral 
            knockin into the TRAC locus of human T cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5632)
  AUTHORS   Blaeschke F, Chen YY, Apathy R, Daniel B, Chen AY, Chen PA, Sandor 
            K, Zhang W, Li Z, Mowery CT, Yamamoto TN, Nyberg WA, To A, Yu R, 
            Bueno R, Kim MC, Schmidt R, Goodman DB, Feuchtinger T, Eyquem J, 
            Jimmie Ye C, Carnevale J, Satpathy AT, Shifrut E, Roth TL, Marson A
  TITLE     Modular pooled discovery of synthetic knockin sequences to program 
            durable cell therapies.
  JOURNAL   Cell. 2023 Sep 14;186(19):4216-4234.e33. doi: 
            10.1016/j.cell.2023.08.013.
  PUBMED    37714135
REFERENCE   2  (bases 1 to 5632)
  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: 
            "10.1016/j.cell.2023.08"; journalName: "Cell"; date: "2023-09-14- 
            14"; volume: "186"; issue: "19"; pages: "4216-4234.e33"
COMMENT     Sequence Label: pIG-601_NY-ESO-1_TCR_GFP
FEATURES             Location/Qualifiers
     source          1..5632
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(171..962,963..1031))
                     /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     794..813
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1032..1136)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1204..1222
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1260..1282)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1382..1401
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1595..1617
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1609..1626
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1610..1626
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1992..2051
                     /codon_start=1
                     /product="2A peptide from equine rhinitis A virus 
                     polyprotein"
                     /label=E2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="QCTNYALLKLAGDVESNPGP"
     primer_bind     complement(2091..2112)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     2699..2720
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2802..2855
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3813..3869
                     /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"
     primer_bind     complement(4642..4658)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4642..4658)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4655..4677)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4666..4682
                     /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(4690..4696,4697..4714,4715..4720))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4735..4756
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4873..4890)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5044..5632)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5124..5143)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
       61 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
      121 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
      181 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
      241 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
      301 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
      361 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
      421 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
      481 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
      541 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
      601 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
      661 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
      721 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
      781 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
      841 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      901 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      961 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     1021 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     1081 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     1141 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
     1201 aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac
     1261 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     1321 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat
     1381 gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat accgcacaga
     1441 tgcgtaagga gaaaataccg catcaggcgc cattcaccat tcaggctgcg caactgttgg
     1501 gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct
     1561 gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg
     1621 gccagtgaat tcgagctcgg tacccgggga tttcaggttt ccttgagtgg caggccaggc
     1681 ctggccgtga acgttcactg aaatcatggc ctcttggcca agattgatag cttgtgcctg
     1741 tccctgagtc ccagtccatc acgagcagct ggtttctaag atgctatttc ccgtataaag
     1801 catgagcacg tgcattcgca cgcccacaga gccccgccct tgtccatcac tggcatctgg
     1861 actccagcct gggttggggc aaagagggaa atgagatcat gtcctaaccc tgatcctctt
     1921 gtcccacaga tatccagaac cctgaccctg ccggatcggg tgggactagt ggcggtggtg
     1981 gtagtggtgg tcagtgcacc aactatgcgc tgctgaaact ggcgggcgat gtggaaagca
     2041 acccgggccc gagcaagggc gaagagctgt tcaccggggt ggtgcccatc ctggtcgagc
     2101 tggacggcga cgtaaacggc cacaagttca gcgtgcgcgg cgagggcgag ggcgatgcca
     2161 ccaacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     2221 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     2281 tgaagcgcca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     2341 tcagcttcaa ggacgacggc acctacaaga cccgcgccga ggtgaagttc gagggcgaca
     2401 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     2461 ggcacaagct ggagtacaac ttcaacagcc acaacgtcta tatcaccgcc gacaagcaga
     2521 agaacggcat caaggccaac ttcaagatcc gccacaacgt ggaggacggc agcgtgcagc
     2581 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     2641 accactacct gagcacccag tccgtgctga gcaaagaccc caacgagaag cgcgatcaca
     2701 tggtcctgct ggagttcgtg accgccgccg ggatcactag ggctaagaga ggttcaggat
     2761 ctggtggtgg tagtggcggc ggaaccggtg ctggaagtgg tgagggcagg ggatctctcc
     2821 ttacttgtgg cgacgtggag gagaaccccg gccccatgag catcggcctc ctgtgctgtg
     2881 cagccttgtc tctcctgtgg gcaggtccag tgaatgctgg tgtcactcag accccaaaat
     2941 tccaggtcct gaagacagga cagagcatga cactgcagtg tgcccaggat atgaaccatg
     3001 aatacatgtc ctggtatcga caagacccag gcatggggct gaggctgatt cattactcag
     3061 ttggtgctgg tatcactgac caaggagaag tccccaatgg ctacaatgtc tccagatcaa
     3121 ccacagagga tttcccgctc aggctgctgt cggctgctcc ctcccagaca tctgtgtact
     3181 tctgtgccag cagttacgtc gggaacaccg gggagctgtt ttttggagaa ggctctaggc
     3241 tgaccgtact ggaggacctg aaaaacgtgt tcccacccga ggtcgctgtg tttgagccat
     3301 cagaagcaga gatctcccac acccaaaagg ccacactggt atgcctggcc acaggcttct
     3361 accccgacca cgtggagctg agctggtggg tgaatgggaa ggaggtgcac agtggggtca
     3421 gcacagaccc gcagcccctc aaggagcagc ccgccctcaa tgactccaga tactgcctga
     3481 gcagccgcct gagggtctcg gccaccttct ggcagaaccc ccgcaaccac ttccgctgtc
     3541 aagtccagtt ctacgggctc tcggagaatg acgagtggac ccaggatagg gccaaacccg
     3601 tcacccagat cgtcagcgcc gaggcctggg gtagagcaga ctgtggcttc acctccgagt
     3661 cttaccagca aggggtcctg tctgccacca tcctctatga gatcttgcta gggaaggcca
     3721 ccttgtatgc cgtgctggtc agtgccctcg tgctgatggc tatggtcaag agaaaggatt
     3781 ccagaggccg ggccaagcgg tccggatccg gagccaccaa cttcagcctg ctgaagcagg
     3841 ccggcgacgt ggaggagaac cccggcccca tggagaccct cttgggcctg cttatccttt
     3901 ggctgcagct gcaatgggtg agcagcaaac aggaggtgac gcagattcct gcagctctga
     3961 gtgtcccaga aggagaaaac ttggttctca actgcagttt cactgatagc gctatttaca
     4021 acctccagtg gtttaggcag gaccctggga aaggtctcac atctctgttg cttattcagt
     4081 caagtcagag agagcaaaca agtggaagac ttaatgcctc gctggataaa tcatcaggac
     4141 gtagtacttt atacattgca gcttctcagc ctggtgactc agccacctac ctctgtgctg
     4201 tgaggcccct gtacggagga agctacatac ctacatttgg aagaggaacc agccttattg
     4261 ttcatccgta tatccagaac cctgaccctg cggtgtacca gctgagagac tctaaatcca
     4321 gtgacaagtc tgtctgccta ttcaccgatt ttgattctca aacaaatgtg tcacaaagta
     4381 aggattctga tgtgtatatc acagacaaaa ctgtgctaga catgaggtct atggacttca
     4441 agagcaacag tgctgtggcc tggagcaaca aatctgactt tgcatgtgca aacgccttca
     4501 acaacagcat tattccagaa gacaccttct tccccagccc aggtaagggc agctttggtg
     4561 ccttcgcagg ctgtttcctt gcttcaggaa tggccatcct ctagagtcga cctgcaggca
     4621 tgcaagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     4681 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     4741 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     4801 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     4861 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     4921 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     4981 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     5041 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     5101 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     5161 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     5221 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     5281 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     5341 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     5401 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     5461 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     5521 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     5581 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aa
//
