LOCUS       Exported                8812 bp ds-DNA     circular SYN 04-ABR-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pD188-tet-pLKO.puro_shGAC
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 8812)
  AUTHORS   Marilia Meira Dias
  TITLE     Direct Submission
  JOURNAL   Exported quarta-feira, 4 de abr de 2018 from SnapGene 2.8.3
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8812
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     CDS             169..768
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /note="PuroR"
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     LTR             896..1129
                     /note="3' LTR (Delta-U3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1201..1322
                     /note="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     rep_origin      1362..1497
                     /note="SV40 ori"
                     /note="SV40 origin of replication"
     promoter        complement(1518..1536)
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1546..1562)
                     /note="M13 fwd"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      1704..2159
                     /direction=RIGHT
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        2185..2289
                     /gene="bla"
                     /note="AmpR promoter"
     CDS             2290..3150
                     /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"
     rep_origin      3321..3909
                     /direction=RIGHT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    4197..4218
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding site"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4233..4263
                     /note="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    4271..4287
                     /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)."
     primer_bind     4295..4311
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        4332..4350
                     /note="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4378..4604
                     /note="RSV promoter"
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             4605..4785
                     /note="5' LTR (truncated)"
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    4832..4957
                     /note="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5450..5683
                     /note="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     protein_bind    6249..6267
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6277..6295
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    6302..6320
                     /note="shGAC"
     misc_feature    6327..6345
                     /note="shGAC"
     misc_feature    6392..6509
                     /note="cPPT/CTS"
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        6558..7068
                     /note="hPGK promoter"
                     /note="human phosphoglycerate kinase 1 promoter"
     intron          7077..7649
                     /note="beta-globin intron"
                     /note="intron from rabbit beta-globin gene"
     promoter        7704..7722
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             7733..8350
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /note="TetR"
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESG"
     misc_feature    8388..149
                     /note="IRES"
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
ORIGIN
        1 tgaaggatgc ccagaaggta ccccattgta tgggatctga tctggggcct cggtgcacat
       61 gctttacatg tgtttagtcg aggttaaaaa aacgtctagg ccccccgaac cacggggacg
      121 tggttttcct ttgaaaaaca cgatgataag gatccaccgg agcttaccat gaccgagtac
      181 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
      241 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
      301 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
      361 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
      421 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
      481 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
      541 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
      601 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg
      661 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
      721 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgacg cccgccccac
      781 gacccgcagc gcccgaccga aaggagcgca cgaccccatg catcggtacc tttaagacca
      841 atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa
      901 gggctaattc actcccaacg aagacaagat ctgctttttg cttgtactgg gtctctctgg
      961 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
     1021 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
     1081 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag tagtagttca
     1141 tgtcatctta ttattcagta tttataactt gcaaagaaat gaatatcaga gagtgagagg
     1201 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     1261 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     1321 tatcatgtct ggctctagct atcccgcccc taactccgcc catcccgccc ctaactccgc
     1381 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     1441 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     1501 ggacgtaccc aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt
     1561 acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc
     1621 ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt
     1681 gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     1741 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     1801 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     1861 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     1921 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     1981 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     2041 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     2101 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
     2161 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     2221 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     2281 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     2341 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     2401 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     2461 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     2521 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     2581 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     2641 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     2701 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     2761 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     2821 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     2881 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     2941 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     3001 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     3061 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     3121 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     3181 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     3241 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     3301 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     3361 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     3421 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     3481 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     3541 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     3601 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     3661 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     3721 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     3781 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     3841 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     3901 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     3961 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     4021 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     4081 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
     4141 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
     4201 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
     4261 ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac
     4321 gccaagcgcg caattaaccc tcactaaagg gaacaaaagc tggagctgca agcttaatgt
     4381 agtcttatgc aatactcttg tagtcttgca acatggtaac gatgagttag caacatgcct
     4441 tacaaggaga gaaaaagcac cgtgcatgcc gattggtgga agtaaggtgg tacgatcgtg
     4501 ccttattagg aaggcaacag acgggtctga catggattgg acgaaccact gaattgccgc
     4561 attgcagaga tattgtattt aagtgcctag ctcgatacat aaacgggtct ctctggttag
     4621 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
     4681 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
     4741 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag
     4801 ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga
     4861 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag
     4921 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag
     4981 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca
     5041 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac
     5101 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga
     5161 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga
     5221 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac
     5281 caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt
     5341 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
     5401 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
     5461 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg
     5521 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
     5581 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
     5641 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
     5701 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
     5761 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca
     5821 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag
     5881 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc
     5941 tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt
     6001 ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga
     6061 cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag
     6121 agagagacag agacagatcc attcgattag tgaacggatc tcgacggtat cgatcacgag
     6181 actagcctcg agcggccgca atatttgcat gtcgctatgt gttctgggaa atcaccataa
     6241 acgtgaaatc cctatcagtg atagagactt ataagttccc tatcagtgat agagacaccg
     6301 gcctctgttc tgtcagagtt ctcgagaact ctgacagaac agaggttttt gaattctcga
     6361 cctcgagaca aatggcagta ttcatccaca attttaaaag aaaagggggg attggggggt
     6421 acagtgcagg ggaaagaata gtagacataa tagcaacaga catacaaact aaagaattac
     6481 aaaaacaaat tacaaaaatt caaaattttc gggtttatta cagggacagc agagatccac
     6541 tttggccgcg gctcgagggg gttggggttg cgccttttcc aaggcagccc tgggtttgcg
     6601 cagggacgcg gctgctctgg gcgtggttcc gggaaacgca gcggcgccga ccctgggtct
     6661 cgcacattct tcacgtccgt tcgcagcgtc acccggatct tcgccgctac ccttgtgggc
     6721 cccccggcga cgcttcctgc tccgccccta agtcgggaag gttccttgcg gttcgcggcg
     6781 tgccggacgt gacaaacgga agccgcacgt ctcactagta ccctcgcaga cggacagcgc
     6841 cagggagcaa tggcagcgcg ccgaccgcga tgggctgtgg ccaatagcgg ctgctcagca
     6901 gggcgcgccg agagcagcgg ccgggaaggg gcggtgcggg aggcggggtg tggggcggta
     6961 gtgtgggccc tgttcctgcc cgcgcggtgt tccgcattct gcaagcctcc ggagcgcacg
     7021 tcggcagtcg gctccctcgt tgaccgaatc accgacctct ctccccaggg ggatctgtga
     7081 gtttggggac ccttgattgt tctttctttt tcgctattgt aaaattcatg ttatatggag
     7141 ggggcaaagt tttcagggtg ttgtttagaa tgggaagatg tcccttgtat caccatggac
     7201 cctcatgata attttgtttc tttcactttc tactctgttg acaaccattg tctcctctta
     7261 ttttcttttc attttctgta actttttcgt taaactttag cttgcatttg taacgaattt
     7321 ttaaattcac ttttgtttat ttgtcagatt gtaagtactt tctctaatca cttttttttc
     7381 aaggcaatca gggtatatta tattgtactt cagcacagtt ttagagaaca attgttataa
     7441 ttaaatgata aggtagaata tttctgcata taaattctgg ctggcgtgga aatattctta
     7501 ttggtagaaa caactacatc ctggtcatca tcctgccttt ctctttatgg ttacaatgat
     7561 atacactgtt tgagatgagg ataaaatact ctgagtccaa accgggcccc tctgctaacc
     7621 atgttcatgc cttcttcttt ttcctacagc tcctgggcaa cgtgctggtt attgtgctgt
     7681 ctcatcattt tggcaaagaa ttgtaatacg actcactata gggcgaattg atatgtctag
     7741 attagataaa agtaaagtga ttaacagcgc attagagctg cttaatgagg tcggaatcga
     7801 aggtttaaca acccgtaaac tcgcccagaa gctaggtgta gagcagccta cattgtattg
     7861 gcatgtaaaa aataagcggg ctttgctcga cgccttagcc attgagatgt tagataggca
     7921 ccatactcac ttttgccctt tagaagggga aagctggcaa gattttttac gtaataacgc
     7981 taaaagtttt agatgtgctt tactaagtca tcgcgatgga gcaaaagtac atttaggtac
     8041 acggcctaca gaaaaacagt atgaaactct cgaaaatcaa ttagcctttt tatgccaaca
     8101 aggtttttca ctagagaatg cattatatgc actcagcgct gtggggcatt ttactttagg
     8161 ttgcgtattg gaagatcaag agcatcaagt cgctaaagaa gaaagggaaa cacctactac
     8221 tgatagtatg ccgccattat tacgacaagc tatcgaatta tttgatcacc aaggtgcaga
     8281 gccagccttc ttattcggcc ttgaattgat catatgcgga ttagaaaaac aacttaaatg
     8341 tgaaagtggg tccgcgtaca gcggctcccg ggagttctag ggatctgccc ctctccctcc
     8401 ccccccccta acgttactgg ccgaagccgc ttggaataag gccggtgtgc gtttgtctat
     8461 atgttatttt ccaccatatt gccgtctttt ggcaatgtga gggcccggaa acctggccct
     8521 gtcttcttga cgagcattcc taggggtctt tcccctctcg ccaaaggaat gcaaggtctg
     8581 ttgaatgtcg tgaaggaagc agttcctctg gaagcttctt gaagacaaac aacgtctgta
     8641 gcgacccttt gcaggcagcg gaacccccca cctggcgaca ggtgcctctg cggccaaaag
     8701 ccacgtgtat aagatacacc tgcaaaggcg gcacaacccc agtgccacgt tgtgagttgg
     8761 atagttgtgg aaagagtcaa atggctctcc tcaagcgtat tcaacaaggg gc
//
