LOCUS       sequence-439171-        8218 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 8218)
  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 8218)
  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 P428L
FEATURES             Location/Qualifiers
     source          1..8218
                     /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"
     primer_bind     complement(1738..1761)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2100..2119
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             2188..2787
                     /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(2188..2207)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2684..2704
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             2968..3393
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(3496..3518)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3654..3674)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3659..3988
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3839..3974
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4010..4029
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4110..4127)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4281..4869)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4361..4380)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5040..5831,5832..5900))
                     /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     5663..5682
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5901..6005)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6073..6091
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        6191..6570
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6571..6774
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             6775..6950
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    7013..7212
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             7413..7829
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     7759..7781
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     7797..7813
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        7869..8218
                     /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    7877..7895
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7913..7931
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7949..7967
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7985..8003
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    8021..8039
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    8057..8075
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    8093..8111
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     8157..8181
                     /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 tcatctcaga 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 ggttcgattt 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 tgggctcagg ggcgggctca ggggcggggc gggcgcccga aggtcctccg gaggcccggc
     2101 attctgcacg cttcaaaagc gcacgtctgc cgcgctgttc tcctcttcct catctccggg
     2161 cctttcgacc tgcagcccaa gcttaccatg accgagtaca agcccacggt gcgcctcgcc
     2221 acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc
     2281 gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa
     2341 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc
     2401 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc
     2461 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc
     2521 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg
     2581 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc
     2641 gagcgcgccg gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac
     2701 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg
     2761 tgcatgaccc gcaagcccgg tgcctgacgc ccgccccacg acccgcagcg cccgaccgaa
     2821 aggagcgcac gaccccatgc atcggcgtct cgagatatca gtggtccagg ctctagtttt
     2881 gactcaacaa tatcaccagc tgaagcctat agagtacgag ccatagataa aataaaagat
     2941 tttatttagt ctccagaaaa aggggggaat gaaagacccc acctgtaggt ttggcaagct
     3001 agcttaagta acgccatttt gcaaggcatg gaaaaataca taactgagaa tagagaagtt
     3061 cagatcaagg tcaggaacag atggaacagg gtcgacccta gagaaccatc agatgtttcc
     3121 agggtgcccc aaggacctga aatgaccctg tgccttattt gaactaacca atcagttcgc
     3181 ttctcgcttc tgttcgcgcg cttctgctcc ccgagctcaa taaaagagcc cacaacccct
     3241 cactcggggc gccagtcctc cgattgactg agtcgcccgg gtacccgtgt atccaataaa
     3301 ccctcttgca gttgcatccg acttgtggtc tcgctgttcc ttgggagggt ctcctctgag
     3361 tgattgacta cccgtcagcg ggggtctttc atttgggggc tcgtccggga tcgggagacc
     3421 cctgcccagg gaccaccgac ccaccaccgg gaggtaagct ggctgcctcg cgcgtttcgg
     3481 tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta
     3541 agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg
     3601 gggcgcagcc atgacccagt cacgtagcga tagcggagtg tagatccggc tgtggaatgt
     3661 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     3721 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     3781 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     3841 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     3901 tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg
     3961 cttttttgga ggcctaggct tttgcaaaaa gcttactggc ttaactatgc ggcatcagag
     4021 cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga
     4081 aaataccgca tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     4141 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     4201 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     4261 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     4321 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     4381 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     4441 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     4501 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     4561 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     4621 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     4681 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     4741 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     4801 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     4861 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     4921 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     4981 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     5041 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     5101 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     5161 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     5221 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     5281 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     5341 gttgttgcca ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     5401 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     5461 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     5521 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     5581 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     5641 tcttgcccgg cgtcaacacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     5701 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     5761 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     5821 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     5881 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     5941 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     6001 ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca
     6061 ttaacctata aaaataggcg tatcacgagg ccctttcgtc ttcaagaatt agcttggcca
     6121 ttgcatacgt tgtatccata tcataatatg tacatttata ttggctcatg tccaacatta
     6181 ccgccatgtt gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
     6241 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
     6301 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
     6361 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     6421 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     6481 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     6541 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     6601 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     6661 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     6721 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcaataa
     6781 aagagcccac aacccctcac tcggcgcgcc agtcttccga tagactgcgt cgcccgggta
     6841 cccgtattcc caataaagcc tcttgctgtt tgcatccgaa tcgtggtctc gctgttcctt
     6901 gggagggtct cctctgagtg attgactacc cacgacgggg gtctttcatt tgggggctcg
     6961 tccgggattt ggagacccct gcccagggac caccgaccca ccaccgggag gtaagctggc
     7021 cagcaactta tctgtgtctg tccgattgtc tagtgtctat gtttgatgtt atgcgcctgc
     7081 gtctgtacta gttagctaac tagctctgta tctggcggac ccgtggtgga actgacgagt
     7141 tctgaacacc cggccgcaac cctgggagac gtcccaggga ctttgggggc cgtttttgtg
     7201 gcccgacctg aggaagggag tcgatgtgga atccgacccc gtcaggatat gtggttctgg
     7261 taggagacga gaacctaaaa cagttcccgc ctccgtctga atttttgctt tcggtttgga
     7321 accgaagccg cgcgtcttgt ctgctgcagc gctgcagcat cgttctgtgt tgtctctgtc
     7381 tgactgtgtt tctgtatttg tctgaaaatt agggccagac tgttaccact cccttaagtt
     7441 tgaccttagg tcactggaaa gatgtcgagc ggatcgctca caaccagtcg gtagatgtca
     7501 agaagagacg ttgggttacc ttctgctctg cagaatggcc aacctttaac gtcggatggc
     7561 cgcgagacgg cacctttaac cgagacctca tcacccaggt taagatcaag gtcttttcac
     7621 ctggcccgca tggacaccca gaccaggtcc cctacatcgt gacctgggaa gccttggctt
     7681 ttgacccccc tccctgggtc aagccctttg tacaccctaa gcctccgcct cctcttcctc
     7741 catccgcccc gtctctcccc cttgaacctc ctcgttcgac cccgcctcga tcctcccttt
     7801 atccagccct cactccttct ctaggcgccg gaattgaaga tctcgatgag gccctttcgt
     7861 cttcactcga gtttactccc tatcagtgat agagaacgta tgaagagttt actccctatc
     7921 agtgatagag aacgtatgca gactttactc cctatcagtg atagagaacg tataaggagt
     7981 ttactcccta tcagtgatag agaacgtatg accagtttac tccctatcag tgatagagaa
     8041 cgtatctaca gtttactccc tatcagtgat agagaacgta tatccagttt actccctatc
     8101 agtgatagag aacgtatgtc gaggtaggcg tgtacggtgg gcgcctataa aagcagagct
     8161 cgtttagtga accgtcagat cgcctggagc aattccacaa cacttttgtc ttatactt
//
