LOCUS       Exported                6336 bp ds-DNA     circular SYN 24-JAN-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pAAV_TRE_Rnd_HA_RINGnoNLS_TPR3_PIR_P2A_GFPii
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6336)
  AUTHORS   Don Arnold
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Mar 13, 2025 from SnapGene 3.2.1
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6336
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="AAV2 ITR"
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     promoter        159..473
                     /label=tight TRE promoter
                     /note="tight TRE promoter"
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    167..185
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    203..221
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    238..256
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    274..292
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    310..328
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    345..363
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    381..399
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             493..758
                     /codon_start=1
                     /note="Random FingR"
                     /translation="LEVVVATPTSLLISWRATQEHARYYRITYGETGGNSPVQEFTVPG
                     SKSTATISGLKPGVDYTITVYAVTREPQASSDQSPISINYRTR"
     misc_feature    766..792
                     /note="HA tag"
     CDS             808..933
                     /codon_start=1
                     /note="Mdm2 RING"
                     /translation="CVICQGRPKNGCIVHGKTGHLMSCFTCAKALAAANAPCPVCR"
     CDS             892..915
                     /codon_start=1
                     /note="NLS mutated"
                     /translation="KALAAANA"
     misc_feature    940..1362
                     /note="TPR3"
     CDS             1369..1668
                     /codon_start=1
                     /note="PIR"
                     /translation="KKKLSKSDIMLVQSANVPSNPTQVPVEESGGFGSHHHNQNYCYQV
                     CLTPESAKTDLMFLKPCSPSRSTDAEHNPCGAIVTGYSDQQPDIISNGSILSNEN"
     misc_feature    1684..1740
                     /note="P2A"
     misc_feature    1750..2462
                     /note="eGFP"
     misc_feature    2466..3074
                     /label=WPRE
                     /note="WPRE"
     polyA_signal    3083..3559
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     repeat_region   3599..3739
                     /label=AAV2 ITR
                     /note="AAV2 ITR"
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     rep_origin      3814..4269
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4551..4655
                     /gene="bla"
                     /label=AmpR promoter
                     /note="bla promoter"
     CDS             4656..5516
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="bla"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5687..6275
                     /direction=RIGHT
                     /label=ori
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtctcga gtttactccc tatcagtgat
      181 agagaacgta tgtcgagttt actccctatc agtgatagag aacgatgtcg agtttactcc
      241 ctatcagtga tagagaacgt atgtcgagtt tactccctat cagtgataga gaacgtatgt
      301 cgagtttact ccctatcagt gatagagaac gtatgtcgag tttatcccta tcagtgatag
      361 agaacgtatg tcgagtttac tccctatcag tgatagagaa cgtatgtcga ggtaggcgtg
      421 tacggtggga ggcctatata agcagagctc gtttagtgaa ccgtcagatc gcctggagaa
      481 ttctctagaa tgctcgaagt cgtcgttgcg actccgacca gcctcctaat cagctggcgg
      541 gcgacgcagg agcacgcgcg ctactaccgc atcacctacg gtgaaactgg tggcaatagc
      601 cctgtccagg aattcaccgt gcctggcagc aagtccactg ctaccatcag cggcctgaaa
      661 cctggtgtcg actataccat cacggtgtac gccgtcacgc gcgagcccca ggccagctcc
      721 gaccagtccc cgatctccat caactaccgc accagatcaa ccggttaccc atacgatgtt
      781 ccagattacg ctggtggaag cacgcgttgt gtgatttgcc aggggcggcc taaaaatggt
      841 tgcattgttc acggcaaaac cggacacctg atgtcatgtt tcacgtgtgc aaaggcacta
      901 gctgcggcga atgcgccctg ccccgtgtgc agagctagca agctctcagg tggcggaggg
      961 agcggcggag gtggctctgg aggtggtggc tcagctgaag catggtacaa tctgggaaac
     1021 gcttattaca agcagggcga ctatcaaaaa gccattgagt attaccaaaa ggctcttgag
     1081 ctagacccta ataacgccga agcgtggtat aacctaggca acgcatatta caagcaggga
     1141 gactaccaaa aggcaatcga atattaccag aaggcactcg aactggaccc aaataacgca
     1201 gaggcttggt ataatctggg aaacgcctac tataaacagg gtgactacca aaaggcgata
     1261 gaggactatc agaaggcctt ggagctcgat ccgaacaatc tgcaacgtag tgctggtggc
     1321 ggagggtctg gtggaggagg ctcaggagga ggtggcgcat caggtaccaa gaagaaactc
     1381 agcaagtcag acatcatgtt ggtgcagagc gccaatgtcc ctagcaaccc gacccaggtg
     1441 ccggtagagg agtccggagg cttcggttcc catcaccaca atcagaacta ttgctatcag
     1501 gtctgcctca ccccagagtc cgccaagacc gacctgatgt tcttgaagcc ctgcagccct
     1561 tcccggagta cagacgctga gcacaatccc tgtggagcca tcgtcaccgg gtacagcgac
     1621 cagcaaccag acatcatttc caacggaagc attttgtcca acgagaatgg tggatcatgt
     1681 acagctacta acttcagcct gctgaagcag gctggagacg tggaggagaa ccctggacct
     1741 ggaggtactg gggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc
     1801 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat
     1861 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc
     1921 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac
     1981 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc
     2041 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc
     2101 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc
     2161 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag
     2221 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg
     2281 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc
     2341 gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat
     2401 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg
     2461 taaaagctta tcgataatca acctctggat tacaaaattt gtgaaagatt gactggtatt
     2521 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
     2581 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct
     2641 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct
     2701 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc
     2761 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
     2821 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc
     2881 tttccttggc tgctcgccta tgttgccacc tggattctgc gcgggacgtc cttctgctac
     2941 gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg
     3001 cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc
     3061 ccgcatcgat accgagatct acgggtggca tccctgtgac ccctccccag tgcctctcct
     3121 ggccctggaa gttgccactc cagtgcccac cagccttgtc ctaataaaat taagttgcat
     3181 cattttgtct gactaggtgt ccttctataa tattatgggg tggagggggg tggtatggag
     3241 caaggggcaa gttgggaaga caacctgtag ggcctgcggg gtctattggg aaccaagctg
     3301 gagtgcagtg gcacaatctt ggctcactgc aatctccgcc tcctgggttc aagcgattct
     3361 cctgcctcag cctcccgagt tgttgggatt ccaggcatgc atgaccaggc tcagctaatt
     3421 tttgtttttt tggtagagac ggggtttcac catattggcc aggctggtct ccaactccta
     3481 atctcaggtg atctacccac cttggcctcc caaattgctg ggattacagg cgtgaaccac
     3541 tgctcccttc cctgtccttc tgattttgta ggtaaccacg tgcggaccga gcggccgcag
     3601 gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc
     3661 gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga
     3721 gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca tctgtgcggt
     3781 atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg
     3841 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     3901 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     3961 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     4021 aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     4081 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     4141 tcaaccctat ctcgggctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     4201 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt
     4261 ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc
     4321 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     4381 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     4441 caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     4501 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     4561 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     4621 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     4681 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     4741 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     4801 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     4861 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     4921 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     4981 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     5041 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     5101 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     5161 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     5221 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     5281 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     5341 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     5401 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     5461 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     5521 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     5581 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     5641 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     5701 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     5761 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     5821 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     5881 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     5941 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     6001 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     6061 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     6121 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     6181 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     6241 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     6301 ggttcctggc cttttgctgg ccttttgctc acatgt
//
