LOCUS       Exported                9044 bp ds-DNA     circular SYN 01-MAR-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pLVX TET3G dd mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9044)
  AUTHORS   Francesco Aulicino
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, March 1, 2018 from SnapGene Viewer 3.3.3
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9044
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        1..350
                     /note="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    9..27
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    45..63
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    81..99
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    117..135
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    153..171
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    189..207
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    225..243
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    394..870
                     /note="dd"
                     /note="DHFR derived, TMP tunable destabilisation domain. 
                     Sui et al. 2014 (Stem Cell Reports. 2014 May 6; 2(5): 
                     721–733.)"
     CDS             871..1578
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /note="mCherry"
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     promoter        1624..2123
                     /note="PGK promoter"
                     /note="mouse phosphoglycerate kinase 1 promoter"
     CDS             2144..2743
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /note="PuroR"
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     misc_feature    2761..3349
                     /note="WPRE"
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             complement(3232..3243)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /note="Factor Xa site"
                     /translation="IEGR"
     LTR             3557..4190
                     /note="3' LTR"
                     /note="3' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(4319..4335)
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4343..4359
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(4367..4397)
                     /note="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    4412..4433
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding site"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4721..5306)
                     /direction=LEFT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5477..6337)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6338..6442)
                     /gene="bla"
                     /note="AmpR promoter"
     polyA_signal    6490..6624
                     /note="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     LTR             6840..7473
                     /note="3' LTR"
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    7520..7645
                     /note="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    8142..8375
                     /note="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    8867..8982
                     /note="cPPT/CTS"
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
ORIGIN
        1 gagtttactc cctatcagtg atagagaacg tatgaagagt ttactcccta tcagtgatag
       61 agaacgtatg cagactttac tccctatcag tgatagagaa cgtataagga gtttactccc
      121 tatcagtgat agagaacgta tgaccagttt actccctatc agtgatagag aacgtatcta
      181 cagtttactc cctatcagtg atagagaacg tatatccagt ttactcccta tcagtgatag
      241 agaacgtatg tcgaggtagg cgtgtacggt gggcgcctat aaaagcagag ctcgtttagt
      301 gaaccgtcag atcgcctgga gcaattccac aacacttttg tcttatactt ggatccgggc
      361 ccgcggccgc cccccgcgga tcccccgggc cacatgatca gcctgatcgc cgccctggcc
      421 gtggactacg tgatcggcat ggagaacgcc atgccctgga acctgcccgc cgacctggcc
      481 tggttcaagc gcaacaccct gaacaagccc gtgatcatgg gccgccacac ctgggagagc
      541 atcggccgcc ccctgcccgg ccgcaagaac atcatcctga gcagccagcc cggcaccgac
      601 gaccgcgtga cctgggtgaa gagcgtggac gaggccatcg ccgcctgcgg cgacgtgccc
      661 gagatcatgg tgatcggcgg cggccgcgtg atcgagcagt tcctgcccaa ggcccagaag
      721 ctgtacctga cccacatcga cgccgaggtg gagggcgaca cccacttccc cgactacgag
      781 cccgacgact gggagagcgt gttcagcgag ttccacgacg ccgacgccca gaacagccac
      841 agctactgct tcgagatcct ggagcgccgc gtgagcaagg gcgaggagga taacatggcc
      901 atcatcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtgaa cggccacgag
      961 ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg
     1021 aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctgtcccc tcagttcatg
     1081 tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt gaagctgtcc
     1141 ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc
     1201 gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa gctgcgcggc
     1261 accaacttcc cctccgacgg ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc
     1321 tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagca gaggctgaag
     1381 ctgaaggacg gcggccacta cgacgctgag gtcaagacca cctacaaggc caagaagccc
     1441 gtgcagctgc ccggcgccta caacgtcaac atcaagttgg acatcacctc ccacaacgag
     1501 gactacacca tcgtggaaca gtacgaacgc gccgagggcc gccactccac cggcggcatg
     1561 gacgagctgt acaagtaacc cggggaattc ctcgagaagg gacgcgtcat atggaattct
     1621 accgggtagg ggaggcgctt ttcccaaggc agtctggagc atgcgcttta gcagccccgc
     1681 tgggcacttg gcgctacaca agtggcctct ggcctcgcac acattccaca tccaccggta
     1741 ggcgccaacc ggctccgttc tttggtggcc ccttcgcgcc accttctact cctcccctag
     1801 tcaggaagtt cccccccgcc ccgcagctcg cgtcgtgcag gacgtgacaa atggaagtag
     1861 cacgtctcac tagtctcgtg cagatggaca gcaccgctga gcaatggaag cgggtaggcc
     1921 tttggggcag cggccaatag cagctttgct ccttcgcttt ctgggctcag aggctgggaa
     1981 ggggtgggtc cgggggcggg ctcaggggcg ggctcagggg cggggcgggc gcccgaaggt
     2041 cctccggagg cccggcattc tgcacgcttc aaaagcgcac gtctgccgcg ctgttctcct
     2101 cttcctcatc tccgggcctt tcgacctgca gcccaagctt accatgaccg agtacaagcc
     2161 cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc
     2221 gttcgccgac taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt
     2281 caccgagctg caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt
     2341 cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc
     2401 ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca
     2461 gcaacagatg gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc
     2521 caccgtcggc gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc
     2581 cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg
     2641 caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga
     2701 aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgacggggcg cgtctggaac
     2761 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     2821 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     2881 atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg
     2941 tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact
     3001 ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct
     3061 attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
     3121 ttgggcactg acaattccgt ggtgttgtcg gggaagctga cgtcctttcc atggctgctc
     3181 gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc
     3241 aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt
     3301 cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcc tggaattaat
     3361 tctgcagtcg agacctagaa aaacatggag caatcacaag tagcaataca gcagctacca
     3421 atgctgattg tgcctggcta gaagcacaag aggaggagga ggtgggtttt tccagtcaca
     3481 cctcaggtac ctttaagacc aatgacttac aaggcagctg tagatcttag ccacttttta
     3541 aaagaaaaga ggggactgga agggctaatt cactcccaac gaagacaaga tatccttgat
     3601 ctgtggatct accacacaca aggctacttc cctgattagc agaactacac accagggcca
     3661 ggggtcagat atccactgac ctttggatgg tgctacaagc tagtaccagt tgagccagat
     3721 aaggtagaag aggccaataa aggagagaac accagcttgt tacaccctgt gagcctgcat
     3781 gggatggatg acccggagag agaagtgtta gagtggaggt ttgacagccg cctagcattt
     3841 catcacgtgg cccgagagct gcatccggag tacttcaaga actgctgata tcgagcttgc
     3901 tacaagggac tttccgctgg ggactttcca gggaggcgtg gcctgggcgg gactggggag
     3961 tggcgagccc tcagatcctg catataagca gctgcttttt gcctgtactg ggtctctctg
     4021 gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc
     4081 tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg
     4141 taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca gtagtagttc
     4201 atgtcatctt attattcagt atttataact tgcaaagaaa tgaatatcag agagtgagag
     4261 gccttgacat tgctagcgtt ttaccgtcga cctctagcta gagcttggcg taatcatggt
     4321 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     4381 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     4441 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     4501 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     4561 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4621 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     4681 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     4741 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     4801 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     4861 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     4921 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     4981 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     5041 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     5101 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     5161 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     5221 gctcttgatc cggcaaacaa accaccgctg gtagcggttt ttttgtttgc aagcagcaga
     5281 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     5341 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5401 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5461 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5521 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5581 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     5641 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     5701 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     5761 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     5821 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     5881 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     5941 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6001 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6061 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     6121 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6181 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6241 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6301 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6361 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6421 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
     6481 cgggagatca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     6541 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     6601 aatgtatctt atcatgtctg gatcaactgg ataactcaag ctaaccaaaa tcatcccaaa
     6661 cttcccaccc cataccctat taccactgcc aattacctag tggtttcatt tactctaaac
     6721 ctgtgattcc tctgaattat tttcatttta aagaaattgt atttgttaaa tatgtactac
     6781 aaacttagta gtttttaaag aaattgtatt tgttaaatat gtactacaaa cttagtagtt
     6841 ggaagggcta attcactccc aaagaagaca agatatcctt gatctgtgga tctaccacac
     6901 acaaggctac ttccctgatt agcagaacta cacaccaggg ccaggggtca gatatccact
     6961 gacctttgga tggtgctaca agctagtacc agttgagcca gataaggtag aagaggccaa
     7021 taaaggagag aacaccagct tgttacaccc tgtgagcctg catgggatgg atgacccgga
     7081 gagagaagtg ttagagtgga ggtttgacag ccgcctagca tttcatcacg tggcccgaga
     7141 gctgcatccg gagtacttca agaactgctg atatcgagct tgctacaagg gactttccgc
     7201 tggggacttt ccagggaggc gtggcctggg cgggactggg gagtggcgag ccctcagatc
     7261 ctgcatataa gcagctgctt tttgcctgta ctgggtctct ctggttagac cagatctgag
     7321 cctgggagct ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt
     7381 gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca
     7441 gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc ccgaacaggg acttgaaagc
     7501 gaaagggaaa ccagaggagc tctctcgacg caggactcgg cttgctgaag cgcgcacggc
     7561 aagaggcgag gggcggcgac tggtgagtac gccaaaaatt ttgactagcg gaggctagaa
     7621 ggagagagat gggtgcgaga gcgtcagtat taagcggggg agaattagat cgcgatggga
     7681 aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa ttaaaacata tagtatgggc
     7741 aagcagggag ctagaacgat tcgcagttaa tcctggcctg ttagaaacat cagaaggctg
     7801 tagacaaata ctgggacagc tacaaccatc ccttcagaca ggatcagaag aacttagatc
     7861 attatataat acagtagcaa ccctctattg tgtgcatcaa aggatagaga taaaagacac
     7921 caaggaagct ttagacaaga tagaggaaga gcaaaacaaa agtaagacca ccgcacagca
     7981 agcggccggc cgctgatctt cagacctgga ggaggagata tgagggacaa ttggagaagt
     8041 gaattatata aatataaagt agtaaaaatt gaaccattag gagtagcacc caccaaggca
     8101 aagagaagag tggtgcagag agaaaaaaga gcagtgggaa taggagcttt gttccttggg
     8161 ttcttgggag cagcaggaag cactatgggc gcagcgtcaa tgacgctgac ggtacaggcc
     8221 agacaattat tgtctggtat agtgcagcag cagaacaatt tgctgagggc tattgaggcg
     8281 caacagcatc tgttgcaact cacagtctgg ggcatcaagc agctccaggc aagaatcctg
     8341 gctgtggaaa gatacctaaa ggatcaacag ctcctgggga tttggggttg ctctggaaaa
     8401 ctcatttgca ccactgctgt gccttggaat gctagttgga gtaataaatc tctggaacag
     8461 atttggaatc acacgacctg gatggagtgg gacagagaaa ttaacaatta cacaagctta
     8521 atacactcct taattgaaga atcgcaaaac cagcaagaaa agaatgaaca agaattattg
     8581 gaattagata aatgggcaag tttgtggaat tggtttaaca taacaaattg gctgtggtat
     8641 ataaaattat tcataatgat agtaggaggc ttggtaggtt taagaatagt ttttgctgta
     8701 ctttctatag tgaatagagt taggcaggga tattcaccat tatcgtttca gacccacctc
     8761 ccaaccccga ggggacccga caggcccgaa ggaatagaag aagaaggtgg agagagagac
     8821 agagacagat ccattcgatt agtgaacgga tctcgacggt atcgccttta aaagaaaagg
     8881 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
     8941 aactaaagaa ctacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
     9001 cagcagagat ccagtttatc gatgaggccc tttcgtcttc actc
//
