LOCUS       sequence-392269-        8083 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Stable and dox. inducible expression of Trim69 Full length, isoform 
            A (codon optimized) after retroviral-mediated gene transfer.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8083)
  AUTHORS   Song Y, Nguyen XN, Kumar A, da Silva C, Picard L, Etienne L, 
            Cimarelli A
  TITLE     Trim69 is a microtubule regulator that acts as a pantropic viral 
            inhibitor.
  JOURNAL   Proc Natl Acad Sci U S A. 2022 Oct 25;119(43):e2211467119. doi: 
            10.1073/pnas.2211467119. Epub 2022 Oct 17.
  PUBMED    36251989
REFERENCE   2  (bases 1 to 8083)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2022-10-25"; volume: "119(43):e2211467119. 
            doi"; pages: " 10.1073/pnas.2211467119. Epub 2022 Oct 17"
COMMENT     Sequence Label: pRetroX-Tight-Pur-Trim69 Full length, isoform ...
FEATURES             Location/Qualifiers
     source          1..8083
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        96..475
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        476..679
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             680..855
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    918..1117
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1318..1734
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1664..1686
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1702..1718
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        1770..2084
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    1778..1796
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1814..1832
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1849..1867
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1885..1903
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1921..1939
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1956..1974
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1992..2010
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2057..2081
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2104..2127
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        3656..4155
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3691..3714)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     4088..4107
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             4176..4775
                     /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(4176..4195)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4672..4692
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4956..5381
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(5484..5506)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5642..5662)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5647..5976
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5827..5962
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5998..6017
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6098..6115)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6269..6857)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6349..6368)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7028..7819,7820..7888))
                     /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     7651..7670
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7889..7993)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8061..8079
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgtcttcaa gaattagctt ggccattgca tacgttgtat ccatatcata atatgtacat
       61 ttatattggc tcatgtccaa cattaccgcc atgttgacat tgattattga ctagttatta
      121 atagtaatca attacggggt cattagttca tagcccatat atggagttcc gcgttacata
      181 acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat
      241 aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga
      301 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc
      361 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt
      421 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat
      481 gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag
      541 tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc
      601 aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga
      661 ggtctatata agcagagctc aataaaagag cccacaaccc ctcactcggc gcgccagtct
      721 tccgatagac tgcgtcgccc gggtacccgt attcccaata aagcctcttg ctgtttgcat
      781 ccgaatcgtg gtctcgctgt tccttgggag ggtctcctct gagtgattga ctacccacga
      841 cgggggtctt tcatttgggg gctcgtccgg gatttggaga cccctgccca gggaccaccg
      901 acccaccacc gggaggtaag ctggccagca acttatctgt gtctgtccga ttgtctagtg
      961 tctatgtttg atgttatgcg cctgcgtctg tactagttag ctaactagct ctgtatctgg
     1021 cggacccgtg gtggaactga cgagttctga acacccggcc gcaaccctgg gagacgtccc
     1081 agggactttg ggggccgttt ttgtggcccg acctgaggaa gggagtcgat gtggaatccg
     1141 accccgtcag gatatgtggt tctggtagga gacgagaacc taaaacagtt cccgcctccg
     1201 tctgaatttt tgctttcggt ttggaaccga agccgcgcgt cttgtctgct gcagcgctgc
     1261 agcatcgttc tgtgttgtct ctgtctgact gtgtttctgt atttgtctga aaattagggc
     1321 cagactgtta ccactccctt aagtttgacc ttaggtcact ggaaagatgt cgagcggatc
     1381 gctcacaacc agtcggtaga tgtcaagaag agacgttggg ttaccttctg ctctgcagaa
     1441 tggccaacct ttaacgtcgg atggccgcga gacggcacct ttaaccgaga cctcatcacc
     1501 caggttaaga tcaaggtctt ttcacctggc ccgcatggac acccagacca ggtcccctac
     1561 atcgtgacct gggaagcctt ggcttttgac ccccctccct gggtcaagcc ctttgtacac
     1621 cctaagcctc cgcctcctct tcctccatcc gccccgtctc tcccccttga acctcctcgt
     1681 tcgaccccgc ctcgatcctc cctttatcca gccctcactc cttctctagg cgccggaatt
     1741 gaagatctga ggccctttcg tcttcactcg agtttactcc ctatcagtga tagagaacgt
     1801 atgtcgagtt tactccctat cagtgataga gaacgatgtc gagtttactc cctatcagtg
     1861 atagagaacg tatgtcgagt ttactcccta tcagtgatag agaacgtatg tcgagtttac
     1921 tccctatcag tgatagagaa cgtatgtcga gtttatccct atcagtgata gagaacgtat
     1981 gtcgagttta ctccctatca gtgatagaga acgtatgtcg aggtaggcgt gtacggtggg
     2041 aggcctatat aagcagagct cgtttagtga accgtcagat cgcctggaga aggatccacc
     2101 atggactaca aagacgatga cgataagggt ggcggagcgg ccgccgaggt gtccaccaac
     2161 cccagctcca acattgaccc tggcgactac gtggagatga acgatagcat cacccacctg
     2221 ccctccaagg tggtcatcca ggacatcacc atggagctgc actgccctct gtgtaacgac
     2281 tggttcagag accctctgat gctgtcctgt ggccacaact tctgcgaggc ctgcatccaa
     2341 gacttttgga ggctgcaggc caaggagacc ttttgccccg agtgcaagat gctgtgccag
     2401 tacaacaact gcacattcaa ccccgtcctg gataagctgg tggagaaaat caagaagctg
     2461 cccctcctga aaggccaccc ccagtgtcct gagcacggcg aaaacctcaa actgttctcc
     2521 aagcccgacg gcaaactgat ttgtttccag tgcaaggatg ccaggctgag cgtgggccag
     2581 tccaaggagt ttctgcaaat ctccgacgcc gtgcacttct tcaccgagga gctggccatt
     2641 cagcagggcc agctggagac caccctgaag gagctgcaaa ccctcaggaa catgcagaag
     2701 gaggccatcg ccgctcacaa ggaaaacaag ctgcacctcc agcagcacgt gtccatggag
     2761 ttcctcaagc tccatcagtt cctgcactcc aaggaaaagg atatcctcac cgagctgaga
     2821 gaggaaggca aagccctgaa tgaagagatg gagctgaacc tgagccagct ccaggagcag
     2881 tgcctgctgg ccaaggacat gctcgtgagc atccaggcca agaccgaaca gcagaacagc
     2941 ttcgacttcc tcaaggatat taccaccctg ctgcactccc tggagcaggg catgaaggtc
     3001 ctggccacca gagagctcat ttccaggaag ctgaacctgg gccagtataa gggccccatt
     3061 cagtacatgg tgtggaggga gatgcaggac accctgtgtc ccggcctgag ccctctgaca
     3121 ctggacccca aaacagccca ccccaatctg gtgctgtcca agagccagac ctccgtgtgg
     3181 catggcgaca tcaagaagat catgcccgat gaccccgaga ggtttgacag cagcgtcgct
     3241 gtgctgggct ccagaggctt taccagcggc aaatggtact gggaagtgga ggtggccaaa
     3301 aagaccaaat ggacagtggg agtggtgagg gaaagcatta tcaggaaggg cagctgtcct
     3361 ctgacacctg agcagggctt ttggctgctg aggctgagaa accagacaga cctgaaggcc
     3421 ctggacctcc ccagcttcag cctgaccctg acaaacaacc tggacaaggt cggcatctac
     3481 ctggactacg agggaggcca gctgagcttc tataatgcca aaaccatgac acacatctat
     3541 accttttcca acacatttat ggagaagctg tacccttatt tctgtccctg cctgaacgac
     3601 ggcggcgaaa ataaggagcc cctgcacatt ctgcaccccc agtgagaatt ctaccgggta
     3661 ggggaggcgc ttttcccaag gcagtctgga gcatgcgctt tagcagcccc gctgggcact
     3721 tggcgctaca caagtggcct ctggcctcgc acacattcca catccaccgg taggcgccaa
     3781 ccggctccgt tctttggtgg ccccttcgcg ccaccttcta ctcctcccct agtcaggaag
     3841 ttcccccccg ccccgcagct cgcgtcgtgc aggacgtgac aaatggaagt agcacgtctc
     3901 actagtctcg tgcagatgga cagcaccgct gagcaatgga agcgggtagg cctttggggc
     3961 agcggccaat agcagctttg ctccttcgct ttctgggctc agaggctggg aaggggtggg
     4021 tccgggggcg ggctcagggg cgggctcagg ggcggggcgg gcgcccgaag gtcctccgga
     4081 ggcccggcat tctgcacgct tcaaaagcgc acgtctgccg cgctgttctc ctcttcctca
     4141 tctccgggcc tttcgacctg cagcccaagc ttaccatgac cgagtacaag cccacggtgc
     4201 gcctcgccac ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg
     4261 actaccccgc cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc
     4321 tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg
     4381 acggcgccgc ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg
     4441 ccgagatcgg cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga
     4501 tggaaggcct cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg
     4561 gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg
     4621 aggcggccga gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc
     4681 ccttctacga gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc
     4741 gcacctggtg catgacccgc aagcccggtg cctgacgccc gccccacgac ccgcagcgcc
     4801 cgaccgaaag gagcgcacga ccccatgcat cggcgtctcg agatatcagt ggtccaggct
     4861 ctagttttga ctcaacaata tcaccagctg aagcctatag agtacgagcc atagataaaa
     4921 taaaagattt tatttagtct ccagaaaaag gggggaatga aagaccccac ctgtaggttt
     4981 ggcaagctag cttaagtaac gccattttgc aaggcatgga aaaatacata actgagaata
     5041 gagaagttca gatcaaggtc aggaacagat ggaacagggt cgaccctaga gaaccatcag
     5101 atgtttccag ggtgccccaa ggacctgaaa tgaccctgtg ccttatttga actaaccaat
     5161 cagttcgctt ctcgcttctg ttcgcgcgct tctgctcccc gagctcaata aaagagccca
     5221 caacccctca ctcggggcgc cagtcctccg attgactgag tcgcccgggt acccgtgtat
     5281 ccaataaacc ctcttgcagt tgcatccgac ttgtggtctc gctgttcctt gggagggtct
     5341 cctctgagtg attgactacc cgtcagcggg ggtctttcat ttgggggctc gtccgggatc
     5401 gggagacccc tgcccaggga ccaccgaccc accaccggga ggtaagctgg ctgcctcgcg
     5461 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     5521 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     5581 gggtgtcggg gcgcagccat gacccagtca cgtagcgata gcggagtgta gatccggctg
     5641 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     5701 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     5761 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     5821 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     5881 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     5941 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc ttactggctt aactatgcgg
     6001 catcagagca gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg
     6061 taaggagaaa ataccgcatc aggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     6121 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     6181 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     6241 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     6301 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     6361 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     6421 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     6481 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     6541 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     6601 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     6661 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg
     6721 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     6781 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     6841 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     6901 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     6961 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     7021 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     7081 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     7141 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     7201 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     7261 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     7321 tgcgcaacgt tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg tttggtatgg
     7381 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     7441 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     7501 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     7561 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     7621 cgagttgctc ttgcccggcg tcaacacggg ataataccgc gccacatagc agaactttaa
     7681 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     7741 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     7801 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     7861 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     7921 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     7981 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta
     8041 tcatgacatt aacctataaa aataggcgta tcacgaggcc ctt
//
