LOCUS       sequence-439167-        8253 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dox-inducible expression of IRSp53 mutant in mammalian cells by 
            retroviral transduction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8253)
  AUTHORS   Landau LM, Chaudhary N, Tien YC, Rogozinska M, Joshi S, Yao C, 
            Crowley J, Hullahalli K, Campbell IW, Waldor MK, Haigis M, Kagan JC
  TITLE     pLxIS-containing domains are biochemically flexible regulators of 
            interferons and metabolism.
  JOURNAL   Mol Cell. 2024 Jul 11;84(13):2436-2454.e10. doi: 
            10.1016/j.molcel.2024.05.030. Epub 2024 Jun 25.
  PUBMED    38925114
REFERENCE   2  (bases 1 to 8253)
  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.molcel.2024.05.030"; journalName: "Mol Cell"; date: 
            "2024-07-11- 11"; volume: "84"; issue: "13"; pages: "2436-2454.e10"
COMMENT     Sequence Label: pRetroX-TRE3G-IRSp53 S269D
FEATURES             Location/Qualifiers
     source          1..8253
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      7..16
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             16..81
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     promoter        1703..2202
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1738..1761)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2135..2154
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             2223..2822
                     /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(2223..2242)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2719..2739
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3003..3428
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(3531..3553)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3689..3709)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3694..4023
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3874..4009
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4045..4064
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4145..4162)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4316..4904)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4396..4415)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5075..5866,5867..5935))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5698..5717
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5936..6040)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6108..6126
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        6226..6605
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6606..6809
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             6810..6985
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    7048..7247
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             7448..7864
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     7794..7816
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     7832..7848
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        7904..8253
                     /label=TRE3GV promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G, optimized for 
                     retroviral and lentiviral vectors"
     protein_bind    7912..7930
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7948..7966
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7984..8002
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    8020..8038
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    8056..8074
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    8092..8110
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    8128..8146
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     8192..8216
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
ORIGIN
        1 ggatccgcca ccatggacta caaggatcac gacggtgatt acaaggacca cgacatcgac
       61 tacaaggacg acgacgacaa gatgtcgctt tcacgctcgg aggagatgca ccggctcacg
      121 gaaaatgtct acaagaccat catggagcag ttcaacccca gcctccgcaa cttcatcgcc
      181 atgggcaaga actatgagaa agcactggca ggtgtcacct tcgctgccaa aggctatttc
      241 gatgctctgg taaagatggg ggagctggcc agcgagagcc agggctctaa ggaacttggg
      301 gacgtcctct tccagatggc tgaggtgcac cggcagatcc agaaccagtt ggaagagacg
      361 ctgaagtctt ttcacaatga gctgctcacg cagctggagc agaaagtaga actggactcc
      421 aggtatctaa gtgctgcact gaagaaatac caaacggaac agaggagcaa aggggacgcc
      481 ctggacaagt gtcaggctga gctgaagaag ctccgcaaga agagccaagg gagtaagaac
      541 cctcagaagt actcagacaa ggagctgcag tacatcgatg ccatcagcaa taagcagggc
      601 gagctggaga actacgtgtc tgacggctac aagacagcac tcactgagga gcgcaggagg
      661 ttctgcttcc tggtggaaaa gcagtgcgct gtggccaaga actctgctgc ttaccattcc
      721 aagggcaagg agttgctggc ccagaagctg cctctgtggc agcaagcctg tgccgacccc
      781 aacaagatcc cagaccgtgc tgtccagctg atgcaacaga tggccaatag caatggctcc
      841 atccttccaa gtgccctgtc agcttccaag tccaacctgg tcatcgacga tcccattcct
      901 ggagccaagc ccttgccagt gccacctgag ctggcaccat ttgtggggcg gatgtctgct
      961 caggagaatg tacctgtcat gaatggcgtc gcaggcccgg acagcgagga ctacaacccc
     1021 tgggctgacc gaaaggctgc ccagcccaaa tccctgtctc ccccgcagtc tcaaagcaag
     1081 ctgagcgact cgtactccaa cacactcccc gtgcgcaaga gcgtgacgcc gaagaacagc
     1141 tatgccacca ctgagaacaa gacactgcca cgttcaagct ccatggcagc tggcctagaa
     1201 cgtaatggcc gcatgcgggt caaggccatt ttctcccacg cggccggtga caatagcact
     1261 ctgctgagct tcaaggaggg cgacctcatc acgctgctag tgcctgaggc ccgtgacggc
     1321 tggcactatg gggagagtga gaagaccaag atgcggggct ggttcccctt ctcctacacc
     1381 cgggtcctgg acagtgacgg aagtgacaga ttgcatatga gcctgcagca gggcaagagc
     1441 agcagcacag gcaacctcct agacaaggat gacctggccc tcccccctcc tgactacggc
     1501 acgtcctccc gggcattccc cacccagaca gccggcacat tcaagcagag accctacagc
     1561 gtggccgttc ctgccttctc tcagggtctg gatgactacg gggcacggtc tgtgagcagg
     1621 aaccccttcg ccaacgtcca tctgaaacca acagtgacca atgacagatc agcccctctg
     1681 ctcagctgat aagaattcta ccgggtaggg gaggcgcttt tcccaaggca gtctggagca
     1741 tgcgctttag cagccccgct gggcacttgg cgctacacaa gtggcctctg gcctcgcaca
     1801 cattccacat ccaccggtag gcgccaaccg gctccgttct ttggtggccc cttcgcgcca
     1861 ccttctactc ctcccctagt caggaagttc ccccccgccc cgcagctcgc gtcgtgcagg
     1921 acgtgacaaa tggaagtagc acgtctcact agtctcgtgc agatggacag caccgctgag
     1981 caatggaagc gggtaggcct ttggggcagc ggccaatagc agctttgctc cttcgctttc
     2041 tgggctcaga ggctgggaag gggtgggtcc gggggcgggc tcaggggcgg gctcaggggc
     2101 ggggcgggcg cccgaaggtc ctccggaggc ccggcattct gcacgcttca aaagcgcacg
     2161 tctgccgcgc tgttctcctc ttcctcatct ccgggccttt cgacctgcag cccaagctta
     2221 ccatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg
     2281 tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg
     2341 accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg
     2401 acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg
     2461 agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg
     2521 gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg
     2581 agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg
     2641 gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc
     2701 tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg
     2761 ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct
     2821 gacgcccgcc ccacgacccg cagcgcccga ccgaaaggag cgcacgaccc catgcatcgg
     2881 cgtctcgaga tatcagtggt ccaggctcta gttttgactc aacaatatca ccagctgaag
     2941 cctatagagt acgagccata gataaaataa aagattttat ttagtctcca gaaaaagggg
     3001 ggaatgaaag accccacctg taggtttggc aagctagctt aagtaacgcc attttgcaag
     3061 gcatggaaaa atacataact gagaatagag aagttcagat caaggtcagg aacagatgga
     3121 acagggtcga ccctagagaa ccatcagatg tttccagggt gccccaagga cctgaaatga
     3181 ccctgtgcct tatttgaact aaccaatcag ttcgcttctc gcttctgttc gcgcgcttct
     3241 gctccccgag ctcaataaaa gagcccacaa cccctcactc ggggcgccag tcctccgatt
     3301 gactgagtcg cccgggtacc cgtgtatcca ataaaccctc ttgcagttgc atccgacttg
     3361 tggtctcgct gttccttggg agggtctcct ctgagtgatt gactacccgt cagcgggggt
     3421 ctttcatttg ggggctcgtc cgggatcggg agacccctgc ccagggacca ccgacccacc
     3481 accgggaggt aagctggctg cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     3541 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
     3601 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac ccagtcacgt
     3661 agcgatagcg gagtgtagat ccggctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     3721 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     3781 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3841 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     3901 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     3961 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     4021 aaaaagctta ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata
     4081 tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgctcttccg
     4141 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     4201 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     4261 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4321 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     4381 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     4441 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     4501 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     4561 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     4621 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     4681 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     4741 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     4801 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     4861 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     4921 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     4981 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5041 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     5101 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5161 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     5221 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5281 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     5341 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct gcaggcatcg
     5401 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     5461 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     5521 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     5581 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     5641 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca acacgggata
     5701 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     5761 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     5821 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     5881 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     5941 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6001 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     6061 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
     6121 cgaggccctt tcgtcttcaa gaattagctt ggccattgca tacgttgtat ccatatcata
     6181 atatgtacat ttatattggc tcatgtccaa cattaccgcc atgttgacat tgattattga
     6241 ctagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
     6301 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
     6361 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
     6421 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
     6481 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
     6541 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
     6601 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
     6661 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
     6721 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
     6781 tacggtggga ggtctatata agcagagctc aataaaagag cccacaaccc ctcactcggc
     6841 gcgccagtct tccgatagac tgcgtcgccc gggtacccgt attcccaata aagcctcttg
     6901 ctgtttgcat ccgaatcgtg gtctcgctgt tccttgggag ggtctcctct gagtgattga
     6961 ctacccacga cgggggtctt tcatttgggg gctcgtccgg gatttggaga cccctgccca
     7021 gggaccaccg acccaccacc gggaggtaag ctggccagca acttatctgt gtctgtccga
     7081 ttgtctagtg tctatgtttg atgttatgcg cctgcgtctg tactagttag ctaactagct
     7141 ctgtatctgg cggacccgtg gtggaactga cgagttctga acacccggcc gcaaccctgg
     7201 gagacgtccc agggactttg ggggccgttt ttgtggcccg acctgaggaa gggagtcgat
     7261 gtggaatccg accccgtcag gatatgtggt tctggtagga gacgagaacc taaaacagtt
     7321 cccgcctccg tctgaatttt tgctttcggt ttggaaccga agccgcgcgt cttgtctgct
     7381 gcagcgctgc agcatcgttc tgtgttgtct ctgtctgact gtgtttctgt atttgtctga
     7441 aaattagggc cagactgtta ccactccctt aagtttgacc ttaggtcact ggaaagatgt
     7501 cgagcggatc gctcacaacc agtcggtaga tgtcaagaag agacgttggg ttaccttctg
     7561 ctctgcagaa tggccaacct ttaacgtcgg atggccgcga gacggcacct ttaaccgaga
     7621 cctcatcacc caggttaaga tcaaggtctt ttcacctggc ccgcatggac acccagacca
     7681 ggtcccctac atcgtgacct gggaagcctt ggcttttgac ccccctccct gggtcaagcc
     7741 ctttgtacac cctaagcctc cgcctcctct tcctccatcc gccccgtctc tcccccttga
     7801 acctcctcgt tcgaccccgc ctcgatcctc cctttatcca gccctcactc cttctctagg
     7861 cgccggaatt gaagatctcg atgaggccct ttcgtcttca ctcgagttta ctccctatca
     7921 gtgatagaga acgtatgaag agtttactcc ctatcagtga tagagaacgt atgcagactt
     7981 tactccctat cagtgataga gaacgtataa ggagtttact ccctatcagt gatagagaac
     8041 gtatgaccag tttactccct atcagtgata gagaacgtat ctacagttta ctccctatca
     8101 gtgatagaga acgtatatcc agtttactcc ctatcagtga tagagaacgt atgtcgaggt
     8161 aggcgtgtac ggtgggcgcc tataaaagca gagctcgttt agtgaaccgt cagatcgcct
     8221 ggagcaattc cacaacactt ttgtcttata ctt
//
