LOCUS       CMV-rtTA-BG-CMV-        5075 bp    DNA     circular SYN 20-AUG-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 5075)
  AUTHORS   Christopher Johnstone
  TITLE     Direct Submission
  JOURNAL   Exported Aug 21, 2025 from SnapGene 8.0.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5075
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     promoter        11..115
                     /gene="bla"
                     /label=AmpR promoter
     CDS             116..976
                     /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     complement(334..353)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1147..1735
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1636..1655
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   1817..1851
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     protein_bind    1888..1912
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    1926..1946
                     /label=pENTR-Gibson-REV
     enhancer        1959..2262
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2263..2466
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             2492..3238
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     polyA_signal    3393..3448
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     misc_feature    3581..3586
                     /label=XbaI
     protein_bind    complement(3586..3610)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    3612
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     enhancer        3643..3946
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3947..4155
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     promoter        3947..4150
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             4177..4776
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     CDS             4177..4199
                     /codon_start=1
                     /label=Puro
                     /translation="MTEYKPT"
     polyA_signal    4777..5001
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     polyA_signal    4777..4784
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5010..5072
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
ORIGIN
        1 ctataatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
       61 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
      121 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
      181 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
      241 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
      301 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
      361 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
      421 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
      481 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
      541 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
      601 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
      661 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
      721 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
      781 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
      841 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
      901 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
      961 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     1021 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     1081 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     1141 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     1201 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     1261 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     1321 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     1381 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     1441 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     1501 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     1561 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     1621 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     1681 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     1741 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     1801 tcctgcgtta tcttcaccct agaaagatag tctgcgtaaa attgacgcat gtccttactc
     1861 atagcaggtg ccactagtgg agttccgaca agtttgtaca aaaaagcagg ctccaccatg
     1921 ggaaccaatt cagtcgactg gatccggtac aggatccgcg ttacataact tacggtaaat
     1981 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     2041 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     2101 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     2161 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     2221 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     2281 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     2341 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     2401 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     2461 agagctcgtt aactctaaaa agaatgtcga catgagccgg ctggacaaga gcaaagtgat
     2521 caacagcgcc ctggaactgc tgaacggcgt gggcatcgag ggcctgacca cccggaagct
     2581 ggcccagaaa ctgggcgtgg aacagcccac cctgtactgg cacgtgaaga acaagcgggc
     2641 cctgctggac gccctgccca tcgagatgct ggaccggcac cacacccaca gctgccctct
     2701 ggaaggcgag agctggcagg acttcctgcg gaacaacgcc aagagctaca gatgcgccct
     2761 gctgagccac cgggacggcg ccaaagtgca cctgggcacc agacccaccg agaagcagta
     2821 cgagacactg gaaaaccagc tggccttcct gtgccagcag ggcttcagcc tggaaaacgc
     2881 cctgtacgcc ctgagcgccg tgggccactt caccctgggc tgcgtgctgg aagaacagga
     2941 acaccaggtc gccaaagagg aaagagagac acccaccacc gacagcatgc cccccctgct
     3001 gaagcaggcc atcgagctgt tcgacagaca gggcgccgag cccgccttcc tgttcggcct
     3061 ggaactgatc atctgcggcc tcgagaagca gctgaagtgc gagagcggcg gacccaccga
     3121 cgccctggac gacttcgacc tggacatgct gcccgccgat gccctggatg attttgatct
     3181 ggatatgctc cctgctgacg ctctcgatga cttcgatctc gatatgctgc caggctaaac
     3241 gcgtgaattc actcctcagg tgcaggctgc ctatcagaag gtggtggctg gtgtggccaa
     3301 tgccctggct cacaaatacc actgagatct ttttccctct gccaaaaatt atggggacat
     3361 catgaagccc cttgagcatc tgacttctgg ctaataaagg aaatttattt tcattgcaat
     3421 agtgtgttgg aattttttgt gtctctcact cggaaggaca tatgggaggg caaatcattt
     3481 aaaacatcag aatgagtatt tggtttagag tttggcaaca tatgcccata tgctggctgc
     3541 catgaacaaa ggttggctat aaagaggtca tcatcggata tctagaccca gctttcttgt
     3601 acaaagtggt cacctgctat gcgctgatat aggtaggcta ctcgttacat aacttacggt
     3661 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     3721 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     3781 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     3841 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     3901 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     3961 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     4021 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     4081 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     4141 aagcagagct gagctcgttt agtgaaccgt cagatgatga ccgagtacaa gcccacggtg
     4201 cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc
     4261 gactaccccg ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag
     4321 ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac
     4381 gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc
     4441 gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag
     4501 atggaaggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc
     4561 ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg
     4621 gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc
     4681 cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg
     4741 cgcacctggt gcatgacccg caagcccggt gcctgactgt gccttctagt tgccagccat
     4801 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     4861 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     4921 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg
     4981 gggatgcggt gggctctatg gagcttacac atgcgtcaat tttacgcatg attatcttta
     5041 acgtacgtca caatatgatt atctttctag ggtta
//
