LOCUS       sequence-8804-ex        9159 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9159)
  AUTHORS   Pluta K, Luce MJ, Bao L, Agha-Mohammadi S, Reiser J
  TITLE     Tight control of transgene expression by lentivirus vectors 
            containing second-generation tetracycline-responsive promoters.
  JOURNAL   J Gene Med. 2005 Jun . 7(6):803-17.
  PUBMED    15655804
REFERENCE   2  (bases 1 to 9159)
  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: "J Gene Med.
            2005 Jun . 7(6):803-17."
COMMENT     Sequence Label: pNL-EGFP/TREPittdU3
FEATURES             Location/Qualifiers
     source          1..9159
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             377..634
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      1260..1395
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     misc_feature    1673..1906
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2091..2135
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     protein_bind    2438..2456
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2475..2493
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2512..2530
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2549..2567
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2586..2604
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2623..2641
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2660..2678
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2697..2715
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2737..2761
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(2836..2838,2839..2841,2842..3552)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2836..2838,2839..2841,2842..3552)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2881..2902)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3142..3161)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3489..3510
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             3757..3989
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(5631..5648)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5802..6390)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5882..5901)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6561..7352,7353..7421))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7184..7203
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7422..7526)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7594..7612
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7663..7685)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    7771..7909
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     7824..7843
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tggaagggct aatttggtcc caaaaaagac aagagatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tggcagaact acacaccagg gccagggatc agatatccac
      121 tgacctttgg atggtgcttc aagttagtac cagttgaacc agagcaagta gaagaggcca
      181 aataaggaga gaagaacagc ttgttacacc ctatgagcca gcatgggatg gaggacccgg
      241 agggagaagt attagtgtgg aagtttgaca gcctcctagc atttcgtcac atggcccgag
      301 agctgcatcc ggagtactac aaagactgct gacatcgagc tttctacaag ggactttccg
      361 ctggggactt tccagggagg tgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 gctacatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgctca aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagtaaa gccagaggag atctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcggta ttaagcgggg gagaattaga taaatgggaa
      841 aaaattcggt taaggccagg gggaaagaaa caatataaac taaaacatat agtatgggca
      901 agcagggagc tagaacgatt cgcagttaat cctggccttt tagagacatc agaaggctgt
      961 agacaaatac tgggacagct acaaccatcc cttcagacag gatcagaaga acttagatca
     1021 ttatataata caatagcagt cctctattgt gtgcatcaaa ggatagatgt aaaagacacc
     1081 aaggaagcct tagataagat agaggaagag caaaacaaag taagaaaaag gcacagcaag
     1141 cagcagctga cacaggaaac aacagccagg tcagccaaaa ttaccctata gtgcagaacc
     1201 tccaggggca aatggtacat caggccatat cacctagaac tttaaatatg actccgccca
     1261 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     1321 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     1381 gcttttttgg aggcctaggc ttttgcaaat gcataaaaga aaagggggga ttggggggta
     1441 cagtgcaggg gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca
     1501 aaaacaaatt acaaaaattc aaaattttcg ggtttattac agggacagca gagatccaca
     1561 attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac
     1621 ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt
     1681 tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga
     1741 cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg
     1801 ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg
     1861 caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt
     1921 gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat
     1981 ctctggaaca gatttggaat aacatgacct ggatggagtg ggacagagaa attaacaatt
     2041 acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac
     2101 aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt
     2161 ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag
     2221 tttttgctgt actttctgta gtgaatagag ttaggcaggg atattcacca ttatcgtttc
     2281 agacccacct cccaatcccg aggggacccg acaggcccga aggaatagaa gaagaaggtg
     2341 gagagagaga cagagacaga tccattcgat tagtgaacgg atctagttcc tcccaccgta
     2401 cacgcctacc tcgacccatc aagtgccacc tgacgtctcc ctatcagtga tagagaagtc
     2461 gacacgtctc gagctcccta tcagtgatag agaaggtacg tctagaacgt ctccctatca
     2521 gtgatagaga agtcgacacg tctcgagctc cctatcagtg atagagaagg tacgtctaga
     2581 acgtctccct atcagtgata gagaagtcga cacgtctcga gctccctatc agtgatagag
     2641 aaggtacgtc tagaacgtct ccctatcagt gatagagaag tcgacacgtc tcgagctccc
     2701 tatcagtgat agagaaggta ccccctatat aagcagagct cgtttagtga accgtcagat
     2761 cgcctggaga cgccatccac gctgttttga cctccataga agacaccggg accgatccag
     2821 cctggatccc cgcggatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc
     2881 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag
     2941 ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc
     3001 gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac
     3061 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag
     3121 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc
     3181 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc
     3241 aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc
     3301 gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc
     3361 agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg
     3421 ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag
     3481 cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac
     3541 gagctgtaca agtccggact cagatctctc gagacctaga aaaacatgga gcaatcacaa
     3601 gtagcaatac agcagctaac aatgctgctt gtgcctggct agaagcacaa gaggaggaag
     3661 aggtgggttt tccagtcaca cctcaggtac ctttaagacc aatgacttac aaggcagctg
     3721 tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt cactcccaaa
     3781 gaagacaaag ctgctttttg cctgtactgg gtctctctgg ttagaccaga tctgagcctg
     3841 ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt
     3901 gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc
     3961 cttttagtca gtgtggaaaa tctctagcac cccccaggag gtagaggttg cagtgagcca
     4021 agatcgcgcc actgcattcc agcctgggca agaaaacaag actgtctaaa ataataataa
     4081 taagttaagg gtattaaata tatttataca tggaggtcat aaaaatatat atatttgggc
     4141 tgggcgcagt ggctcacacc tgcgcccggc cctttgggag gccgaggcag gtggatcacc
     4201 tgagtttggg agttccagac cagcctgacc aacatggaga aaccccttct ctgtgtattt
     4261 ttagtagatt ttattttatg tgtattttat tcacaggtat ttctggaaaa ctgaaactgt
     4321 ttttcctcta ctctgatacc acaagaatca tcagcacaga ggaagacttc tgtgatcaaa
     4381 tgtggtggga gagggaggtt ttcaccagca catgagcagt cagttctgcc gcagactcgg
     4441 cgggtgtcct tcggttcagt tccaacaccg cctgcctgga gagaggtcag accacagggt
     4501 gagggctcag tccccaagac ataaacaccc aagacataaa cacccaacag gtccaccccg
     4561 cctgctgccc aggcagagcc gattcaccaa gacgggaatt aggatagaga aagagtaagt
     4621 cacacagagc cggctgtgcg ggagaacgga gttctattat gactcaaatc agtctcccca
     4681 agcattcggg gatcagagtt tttaaggata acttagtgtg tagggggcca gtgagttgga
     4741 gatgaaagcg tagggagtcg aaggtgtcct tttgcgccga gtcagttcct gggtgggggc
     4801 cacaagatcg gatgagccag tttatcaatc cgggggtgcc agctgatcca tggagtgcag
     4861 ggtctgcaaa atatctcaag cactgattga tcttaggttt tacaatagtg atgttacccc
     4921 aggaacaatt tggggaaggt cagaatcttg tagcctgtag ctgcatgact cctaaaccat
     4981 aatttctttt ttgttttttt ttttttattt ttgagacagg gtctcactct gtcacctagg
     5041 ctggagtgca gtggtgcaat cacagctcac tgcagcctca acgtcgtaag ctcaagcgat
     5101 cctcccacct cagcctgcct ggtagctgag actacaagcg acgccccagt taatttttgt
     5161 atttttggta gaggcagcgt tttgccgtgt ggccctggct ggtctcgaac tcctgggctc
     5221 aagtgatcca gcctcagcct cccaaagtgc tgggacaacc ggggccagtc actgcacctg
     5281 gccctaaacc ataatttcta atcttttggc taatttgtta gtcctacaaa ggcagtctag
     5341 tccccaggca aaaagggggt ttgtttcggg aaagggctgt tactgtcttt gtttcaaact
     5401 ataaactaag ttcctcctaa acttagttcg gcctacaccc aggaatgaac aaggagagct
     5461 tggaggttag aagcacgatg gaattggtta ggtcagatct ctttcactgt ctgagttata
     5521 attttgcaat ggtggttcaa agactgcccg cttctgacac cagtcgctgc attaatgaat
     5581 cggccaacgc gcggggagag gcggtttgcg tattggcgct cttccgcttc ctcgctcact
     5641 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5701 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5761 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5821 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5881 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5941 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc
     6001 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     6061 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     6121 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     6181 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     6241 aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     6301 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     6361 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     6421 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     6481 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     6541 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     6601 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6661 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     6721 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     6781 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6841 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     6901 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6961 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     7021 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     7081 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     7141 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     7201 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     7261 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     7321 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     7381 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     7441 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     7501 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa
     7561 gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt
     7621 cttcaagaac tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
     7681 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     7741 cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag
     7801 cggagtgtac tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat
     7861 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc
     7921 attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca
     7981 ggggaggcag agattgcagt aagctgagat cgcagcactg cactccagcc tgggcgacag
     8041 agtaagactc tgtctcaaaa ataaaataaa taaatcaatc agatattcca atcttttcct
     8101 ttatttattt atttattttc tattttggaa acacagtcct tccttattcc agaattacac
     8161 atatattcta tttttcttta tatgctccag ttttttttag accttcacct gaaatgtgtg
     8221 tatacaaaat ctaggccagt ccagcagagc ctaaaggtaa aaaataaaat aataaaaaat
     8281 aaataaaatc tagctcactc cttcacatca aaatggagat acagctgtta gcattaaata
     8341 ccaaataacc catcttgtcc tcaataattt taagcgcctc tctccaccac atctaactcc
     8401 tgtcaaaggc atgtgcccct tccgggcgct ctgctgtgct gccaaccaac tggcatgtgg
     8461 actctgcagg gtccctaact gccaagcccc acagtgtgcc ctgaggctgc cccttccttc
     8521 tagcggctgc ccccactcgg ctttgctttc cctagtttca gttacttgcg ttcagccaag
     8581 gtctgaaact aggtgcgcac agagcggtaa gactgcgaga gaaagagacc agctttacag
     8641 ggggtttatc acagtgcacc ctgacagtcg tcagcctcac agggggttta tcacattgca
     8701 ccctgacagt cgtcagcctc acagggggtt tatcacagtg cacccttaca atcattccat
     8761 ttgattcaca atttttttag tctctactgt gcctaacttg taagttaaat ttgatcagag
     8821 gtgtgttccc agaggggaaa acagtatata cagggttcag tactatcgca tttcaggcct
     8881 ccacctgggt cttggaatgt gtcccccgag gggtgatgac tacctcagtt ggatctccac
     8941 aggtcacagt gacacaagat aaccaagaca cctcccaagg ctaccacaat gggccgccct
     9001 ccacgtgcac atggccggag gaactgccat gtcggaggtg caagcacacc tgcgcatcag
     9061 agtccttggt gtggagggag ggaccagcgc agcttccagc catccacctg atgaacagaa
     9121 cctagggaaa gccccagttc tacttacacc aggaaaggc
//
