LOCUS       sequence-394441-        7059 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dox-inducible expression of NINJ1 gene  in mammalian cells .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7059)
  AUTHORS   Borges JP, Saetra RSR, Volchuk A, Bugge M, Devant P, Sporsheim B, 
            Kilburn BR, Evavold CL, Kagan JC, Goldenberg NM, Flo TH, Steinberg 
            BE
  TITLE     Glycine inhibits NINJ1 membrane clustering to suppress plasma 
            membrane rupture in cell death.
  JOURNAL   Elife. 2022 Dec 5;11:e78609. doi: 10.7554/eLife.78609.
  PUBMED    36468682
REFERENCE   2  (bases 1 to 7059)
  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: "Elife."; 
            date: "2022-12-5"; volume: "11:e78609. doi"; pages: " 
            10.7554/eLife.78609"
COMMENT     Sequence Label: pRetroX-TRE3G_NINJ1
FEATURES             Location/Qualifiers
     source          1..7059
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        296..499
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             500..675
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    738..937
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1138..1554
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1484..1506
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1522..1538
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        1594..1943
                     /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    1602..1620
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1638..1656
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1674..1692
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1710..1728
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1746..1764
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1782..1800
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1818..1836
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1882..1906
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1953..1962
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1962..1985
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        2452..2951
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2487..2510)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2884..2903
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             2972..3571
                     /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(2972..2991)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3468..3488
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3752..4177
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(4280..4302)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4438..4458)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4443..4772
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4623..4758
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4794..4813
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4894..4911)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5065..5653)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5145..5164)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5824..6615,6616..6684))
                     /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     6447..6466
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6685..6789)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6857..6875
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        join(6975..7059,1..295)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat
       61 aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga
      121 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc
      181 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt
      241 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat
      301 gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag
      361 tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc
      421 aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga
      481 ggtctatata agcagagctc aataaaagag cccacaaccc ctcactcggc gcgccagtct
      541 tccgatagac tgcgtcgccc gggtacccgt attcccaata aagcctcttg ctgtttgcat
      601 ccgaatcgtg gtctcgctgt tccttgggag ggtctcctct gagtgattga ctacccacga
      661 cgggggtctt tcatttgggg gctcgtccgg gatttggaga cccctgccca gggaccaccg
      721 acccaccacc gggaggtaag ctggccagca acttatctgt gtctgtccga ttgtctagtg
      781 tctatgtttg atgttatgcg cctgcgtctg tactagttag ctaactagct ctgtatctgg
      841 cggacccgtg gtggaactga cgagttctga acacccggcc gcaaccctgg gagacgtccc
      901 agggactttg ggggccgttt ttgtggcccg acctgaggaa gggagtcgat gtggaatccg
      961 accccgtcag gatatgtggt tctggtagga gacgagaacc taaaacagtt cccgcctccg
     1021 tctgaatttt tgctttcggt ttggaaccga agccgcgcgt cttgtctgct gcagcgctgc
     1081 agcatcgttc tgtgttgtct ctgtctgact gtgtttctgt atttgtctga aaattagggc
     1141 cagactgtta ccactccctt aagtttgacc ttaggtcact ggaaagatgt cgagcggatc
     1201 gctcacaacc agtcggtaga tgtcaagaag agacgttggg ttaccttctg ctctgcagaa
     1261 tggccaacct ttaacgtcgg atggccgcga gacggcacct ttaaccgaga cctcatcacc
     1321 caggttaaga tcaaggtctt ttcacctggc ccgcatggac acccagacca ggtcccctac
     1381 atcgtgacct gggaagcctt ggcttttgac ccccctccct gggtcaagcc ctttgtacac
     1441 cctaagcctc cgcctcctct tcctccatcc gccccgtctc tcccccttga acctcctcgt
     1501 tcgaccccgc ctcgatcctc cctttatcca gccctcactc cttctctagg cgccggaatt
     1561 gaagatctcg atgaggccct ttcgtcttca ctcgagttta ctccctatca gtgatagaga
     1621 acgtatgaag agtttactcc ctatcagtga tagagaacgt atgcagactt tactccctat
     1681 cagtgataga gaacgtataa ggagtttact ccctatcagt gatagagaac gtatgaccag
     1741 tttactccct atcagtgata gagaacgtat ctacagttta ctccctatca gtgatagaga
     1801 acgtatatcc agtttactcc ctatcagtga tagagaacgt atgtcgaggt aggcgtgtac
     1861 ggtgggcgcc tataaaagca gagctcgttt agtgaaccgt cagatcgcct ggagcaattc
     1921 cacaacactt ttgtcttata cttggatccg ccgccaccat ggactacaaa gacgatgacg
     1981 acaaggagtc tggcaccgag gaatatgagc tgaacggtga tttgcgccct ggctctcctg
     2041 gttctcctga tgccctccca ccaaggtggg ggcttcggaa tcgccctatc aatgtaaatc
     2101 actacgcaaa taagaaatca gcagcagagt ctatgctgga cattgcactc ttgatggcta
     2161 acgcttctca gttgaaggct gtggtagagc aaggaaacga ttttgcattt tttgtcccct
     2221 tggtggtatt gatttcaata agcctggtac ttcagattgg agtcggggta ctcctgatct
     2281 tcttggtgaa atatgacctc aacaatccag ccaaacacgc aaagctggat tttcttaata
     2341 atctcgcaac cggactggta tttattatcg tagtcgttaa catttttatc accgcatttg
     2401 gggtccagaa gccagtgatg gatgtcgccc caagacaata ggaattctac cgggtagggg
     2461 aggcgctttt cccaaggcag tctggagcat gcgctttagc agccccgctg ggcacttggc
     2521 gctacacaag tggcctctgg cctcgcacac attccacatc caccggtagg cgccaaccgg
     2581 ctccgttctt tggtggcccc ttcgcgccac cttctactcc tcccctagtc aggaagttcc
     2641 cccccgcccc gcagctcgcg tcgtgcagga cgtgacaaat ggaagtagca cgtctcacta
     2701 gtctcgtgca gatggacagc accgctgagc aatggaagcg ggtaggcctt tggggcagcg
     2761 gccaatagca gctttgctcc ttcgctttct gggctcagag gctgggaagg ggtgggtccg
     2821 ggggcgggct caggggcggg ctcaggggcg gggcgggcgc ccgaaggtcc tccggaggcc
     2881 cggcattctg cacgcttcaa aagcgcacgt ctgccgcgct gttctcctct tcctcatctc
     2941 cgggcctttc gacctgcagc ccaagcttac catgaccgag tacaagccca cggtgcgcct
     3001 cgccacccgc gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta
     3061 ccccgccacg cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca
     3121 agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg
     3181 cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga
     3241 gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga
     3301 aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt
     3361 ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc
     3421 ggccgagcgc gccggggtgc ccgccttcct ggagacctcc gcgccccgca acctcccctt
     3481 ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac
     3541 ctggtgcatg acccgcaagc ccggtgcctg acgcccgccc cacgacccgc agcgcccgac
     3601 cgaaaggagc gcacgacccc atgcatcggc gtctcgagat atcagtggtc caggctctag
     3661 ttttgactca acaatatcac cagctgaagc ctatagagta cgagccatag ataaaataaa
     3721 agattttatt tagtctccag aaaaaggggg gaatgaaaga ccccacctgt aggtttggca
     3781 agctagctta agtaacgcca ttttgcaagg catggaaaaa tacataactg agaatagaga
     3841 agttcagatc aaggtcagga acagatggaa cagggtcgac cctagagaac catcagatgt
     3901 ttccagggtg ccccaaggac ctgaaatgac cctgtgcctt atttgaacta accaatcagt
     3961 tcgcttctcg cttctgttcg cgcgcttctg ctccccgagc tcaataaaag agcccacaac
     4021 ccctcactcg gggcgccagt cctccgattg actgagtcgc ccgggtaccc gtgtatccaa
     4081 taaaccctct tgcagttgca tccgacttgt ggtctcgctg ttccttggga gggtctcctc
     4141 tgagtgattg actacccgtc agcgggggtc tttcatttgg gggctcgtcc gggatcggga
     4201 gacccctgcc cagggaccac cgacccacca ccgggaggta agctggctgc ctcgcgcgtt
     4261 tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc
     4321 tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt
     4381 gtcggggcgc agccatgacc cagtcacgta gcgatagcgg agtgtagatc cggctgtgga
     4441 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     4501 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     4561 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     4621 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     4681 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     4741 gaggcttttt tggaggccta ggcttttgca aaaagcttac tggcttaact atgcggcatc
     4801 agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag
     4861 gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4921 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4981 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     5041 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     5101 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     5161 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     5221 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     5281 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     5341 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     5401 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     5461 tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat
     5521 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     5581 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     5641 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     5701 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5761 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5821 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5881 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5941 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     6001 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     6061 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     6121 caacgttgtt gccattgctg caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     6181 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     6241 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     6301 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     6361 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     6421 ttgctcttgc ccggcgtcaa cacgggataa taccgcgcca catagcagaa ctttaaaagt
     6481 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6541 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6601 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6661 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6721 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6781 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     6841 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcaag aattagcttg
     6901 gccattgcat acgttgtatc catatcataa tatgtacatt tatattggct catgtccaac
     6961 attaccgcca tgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc
     7021 attagttcat agcccatata tggagttccg cgttacata
//
