LOCUS       sequence-439195-        7677 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dox-inducible expression of Tir in mammalian cells by retroviral 
            transduction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7677)
  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 7677)
  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-Tir
FEATURES             Location/Qualifiers
     source          1..7677
                     /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             214..237
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        1127..1626
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1162..1185)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     1559..1578
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             1647..2246
                     /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(1647..1666)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2143..2163
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             2427..2852
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(2955..2977)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3113..3133)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3118..3447
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3298..3433
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3469..3488
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3569..3586)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3740..4328)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3820..3839)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4499..5290,5291..5359))
                     /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     5122..5141
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5360..5464)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5532..5550
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        5650..6029
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6030..6233
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             6234..6409
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    6472..6671
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             6872..7288
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     7218..7240
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     7256..7272
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        7328..7677
                     /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    7336..7354
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7372..7390
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7408..7426
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7444..7462
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7480..7498
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7516..7534
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7552..7570
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     7616..7640
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
ORIGIN
        1 ggatccgcca ccatggatcg ggctgtttcc caagtagctc tcgaaaacga tgagagagag
       61 gccaaaaaca catggcgact tatctttcgg attgcaatac ttttcctcac agtagttacc
      121 ttggctattt ctgttgctag tctcctgtat tcaatgggcg catccacccc atccgatctt
      181 gtgggaatac ctactcgaat ctctagggct atggactaca aagacgatga cgacaagacc
      241 cctgaacccg acgaccccac caccacagac cctgatcaag cagccaatgc agccgaatca
      301 gccaccaaag accaattgac ccaagaagca tttaagaatc ccgaaaacca aaaggtgaac
      361 attgatgcta atggtaatgc tattccaagt ggcgagttga aagatgatat cgtggagcaa
      421 attgcccaac aggccaaaga agcaggcgaa gtggcaagac agcaggctgt tgagtctaac
      481 gcccaagccc aacaacgcta tgaggatcaa cacgctcgca ggcaagagga gctgcaactt
      541 agctctggaa tcggatatgg cctctcctct gctctgatag tggctggtgg cataggtgct
      601 ggcgtcacca ccgctttgca tcgccggaat cagcccgctg agcaaactac cactacaacc
      661 acacacaccg tggtccaaca acaaactggt ggcaacaccc ctgcacaagg cggcacagat
      721 gctacacgag ctgaggacgc atccctgaat cgcagggaca gtcaaggttc cgtggcaagt
      781 acacactggt ctgatagcag ctccgaagtg gttaatccct acgctgaagt cgggggcgct
      841 agaaattctc ttagcgcaca tcaacctgaa gaacacattt atgacgaggt cgctgcagat
      901 ccaggatatt cagtgataca aaatttcagc gggtctggtc ccgtcactgg tagactcata
      961 ggaacacctg gtcaagggat tcagagcacc tacgcacttc tcgctaatag cggagggctc
     1021 cgacttggaa tgggcggcct gacttctgga ggagagagcg cagtttcctc cgtaaacgct
     1081 gcccccactc cagggccagt tcggttcgtt tgataagaat tctaccgggt aggggaggcg
     1141 cttttcccaa ggcagtctgg agcatgcgct ttagcagccc cgctgggcac ttggcgctac
     1201 acaagtggcc tctggcctcg cacacattcc acatccaccg gtaggcgcca accggctccg
     1261 ttctttggtg gccccttcgc gccaccttct actcctcccc tagtcaggaa gttccccccc
     1321 gccccgcagc tcgcgtcgtg caggacgtga caaatggaag tagcacgtct cactagtctc
     1381 gtgcagatgg acagcaccgc tgagcaatgg aagcgggtag gcctttgggg cagcggccaa
     1441 tagcagcttt gctccttcgc tttctgggct cagaggctgg gaaggggtgg gtccgggggc
     1501 gggctcaggg gcgggctcag gggcggggcg ggcgcccgaa ggtcctccgg aggcccggca
     1561 ttctgcacgc ttcaaaagcg cacgtctgcc gcgctgttct cctcttcctc atctccgggc
     1621 ctttcgacct gcagcccaag cttaccatga ccgagtacaa gcccacggtg cgcctcgcca
     1681 cccgcgacga cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg
     1741 ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac
     1801 tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg
     1861 cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg
     1921 gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag atggaaggcc
     1981 tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc
     2041 ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg
     2101 agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc cccttctacg
     2161 agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt
     2221 gcatgacccg caagcccggt gcctgacgcc cgccccacga cccgcagcgc ccgaccgaaa
     2281 ggagcgcacg accccatgca tcggcgtctc gagatatcag tggtccaggc tctagttttg
     2341 actcaacaat atcaccagct gaagcctata gagtacgagc catagataaa ataaaagatt
     2401 ttatttagtc tccagaaaaa ggggggaatg aaagacccca cctgtaggtt tggcaagcta
     2461 gcttaagtaa cgccattttg caaggcatgg aaaaatacat aactgagaat agagaagttc
     2521 agatcaaggt caggaacaga tggaacaggg tcgaccctag agaaccatca gatgtttcca
     2581 gggtgcccca aggacctgaa atgaccctgt gccttatttg aactaaccaa tcagttcgct
     2641 tctcgcttct gttcgcgcgc ttctgctccc cgagctcaat aaaagagccc acaacccctc
     2701 actcggggcg ccagtcctcc gattgactga gtcgcccggg tacccgtgta tccaataaac
     2761 cctcttgcag ttgcatccga cttgtggtct cgctgttcct tgggagggtc tcctctgagt
     2821 gattgactac ccgtcagcgg gggtctttca tttgggggct cgtccgggat cgggagaccc
     2881 ctgcccaggg accaccgacc caccaccggg aggtaagctg gctgcctcgc gcgtttcggt
     2941 gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa
     3001 gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg
     3061 ggcgcagcca tgacccagtc acgtagcgat agcggagtgt agatccggct gtggaatgtg
     3121 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     3181 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     3241 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     3301 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     3361 atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc
     3421 ttttttggag gcctaggctt ttgcaaaaag cttactggct taactatgcg gcatcagagc
     3481 agattgtact gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa
     3541 aataccgcat caggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     3601 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     3661 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     3721 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     3781 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     3841 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     3901 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     3961 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     4021 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     4081 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     4141 agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg
     4201 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     4261 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     4321 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     4381 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     4441 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     4501 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     4561 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     4621 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     4681 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     4741 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     4801 ttgttgccat tgctgcaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     4861 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     4921 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     4981 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     5041 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     5101 cttgcccggc gtcaacacgg gataataccg cgccacatag cagaacttta aaagtgctca
     5161 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     5221 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     5281 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     5341 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     5401 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     5461 cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat
     5521 taacctataa aaataggcgt atcacgaggc cctttcgtct tcaagaatta gcttggccat
     5581 tgcatacgtt gtatccatat cataatatgt acatttatat tggctcatgt ccaacattac
     5641 cgccatgttg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
     5701 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
     5761 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
     5821 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
     5881 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
     5941 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
     6001 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
     6061 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
     6121 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
     6181 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcaataaa
     6241 agagcccaca acccctcact cggcgcgcca gtcttccgat agactgcgtc gcccgggtac
     6301 ccgtattccc aataaagcct cttgctgttt gcatccgaat cgtggtctcg ctgttccttg
     6361 ggagggtctc ctctgagtga ttgactaccc acgacggggg tctttcattt gggggctcgt
     6421 ccgggatttg gagacccctg cccagggacc accgacccac caccgggagg taagctggcc
     6481 agcaacttat ctgtgtctgt ccgattgtct agtgtctatg tttgatgtta tgcgcctgcg
     6541 tctgtactag ttagctaact agctctgtat ctggcggacc cgtggtggaa ctgacgagtt
     6601 ctgaacaccc ggccgcaacc ctgggagacg tcccagggac tttgggggcc gtttttgtgg
     6661 cccgacctga ggaagggagt cgatgtggaa tccgaccccg tcaggatatg tggttctggt
     6721 aggagacgag aacctaaaac agttcccgcc tccgtctgaa tttttgcttt cggtttggaa
     6781 ccgaagccgc gcgtcttgtc tgctgcagcg ctgcagcatc gttctgtgtt gtctctgtct
     6841 gactgtgttt ctgtatttgt ctgaaaatta gggccagact gttaccactc ccttaagttt
     6901 gaccttaggt cactggaaag atgtcgagcg gatcgctcac aaccagtcgg tagatgtcaa
     6961 gaagagacgt tgggttacct tctgctctgc agaatggcca acctttaacg tcggatggcc
     7021 gcgagacggc acctttaacc gagacctcat cacccaggtt aagatcaagg tcttttcacc
     7081 tggcccgcat ggacacccag accaggtccc ctacatcgtg acctgggaag ccttggcttt
     7141 tgacccccct ccctgggtca agccctttgt acaccctaag cctccgcctc ctcttcctcc
     7201 atccgccccg tctctccccc ttgaacctcc tcgttcgacc ccgcctcgat cctcccttta
     7261 tccagccctc actccttctc taggcgccgg aattgaagat ctcgatgagg ccctttcgtc
     7321 ttcactcgag tttactccct atcagtgata gagaacgtat gaagagttta ctccctatca
     7381 gtgatagaga acgtatgcag actttactcc ctatcagtga tagagaacgt ataaggagtt
     7441 tactccctat cagtgataga gaacgtatga ccagtttact ccctatcagt gatagagaac
     7501 gtatctacag tttactccct atcagtgata gagaacgtat atccagttta ctccctatca
     7561 gtgatagaga acgtatgtcg aggtaggcgt gtacggtggg cgcctataaa agcagagctc
     7621 gtttagtgaa ccgtcagatc gcctggagca attccacaac acttttgtct tatactt
//
