LOCUS       V011750                 8880 bp    DNA     circular SYN 13-MAY-2021
DEFINITION  Exported.
ACCESSION   V011750
VERSION     V011750
KEYWORDS    tet-pLKO-sgRNA-puro
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 8880)
  AUTHORS   Huang HT, Seo HS, Zhang T, Wang Y, Jiang B, Li Q, Buckley DL, Nabet
            B, Roberts JM, Paulk J, Dastjerdi S, Winter GE, McLauchlan H, Moran
            J, Bradner JE, Eck MJ, Dhe-Paganon S, Zhao JJ, Gray NS
  TITLE     MELK is not necessary for the proliferation of basal-like breast
            cancer cells.
  JOURNAL   Elife. 2017 Sep 19;6. pii: e26693. doi: 10.7554/eLife.26693.
   PUBMED   28926338
REFERENCE   2  (bases 1 to 8880)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 8880)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Elife.";
            date: "2017-09-19"; pages: "
            10.7554/eLife.26693"
            SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8880
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        7..507
                     /label="hPGK promoter"
                     /note="human phosphoglycerate kinase 1 promoter"
     intron          516..1088
                     /label="beta-globin intron"
                     /note="intron from rabbit beta-globin gene"
     primer_bind     1096..1115
                     /label="pCAG-F"
                     /note="Rabbit beta-globin intron, for pCAG plasmids,
                     forward primer"
     promoter        1143..1161
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1172..1792
                     /label="TetR"
                     /note="tetracycline repressor TetR"
     misc_feature    1827..2400
                     /label="IRES"
                     /note="internal ribosome entry site (IRES) of the
                     encephalomyocarditis virus (EMCV)"
     CDS             2420..3016
                     /label="PuroR"
                     /note="puromycin N-acetyltransferase"
     LTR             3147..3380
                     /label="3' LTR (Delta-U3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3452..3586
                     /label="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     rep_origin      3613..3748
                     /label="SV40 ori"
                     /note="SV40 origin of replication"
     promoter        complement(3769..3787)
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3797..3813)
                     /label="M13 fwd"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     rep_origin      3955..4410
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        4436..4540
                     /label="AmpR promoter"
     CDS             4541..5398
                     /label="AmpR"
                     /note="beta-lactamase"
     rep_origin      5572..6160
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     primer_bind     6314..6331
                     /label="L4440"
                     /note="L4440 vector, forward primer"
     protein_bind    6448..6469
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        6484..6514
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    6522..6538
                     /label="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     6546..6562
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     promoter        6583..6601
                     /label="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6629..6855
                     /label="RSV promoter"
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6856..7036
                     /label="5' LTR (truncated)"
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    7083..7208
                     /label="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    7701..7934
                     /label="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
     CDS             8119..8163
                     /label="gp41 peptide"
                     /note="antigenic peptide corresponding to amino acids 655
                     to 669 of the HIV envelope protein gp41 (Lutje Hulsik et
                     al., 2013)"
     CDS             8312..8353
                     /note="Protein Tat from Human immunodeficiency virus type 1
                     group M subtype B (isolate WMJ22). Accession#: P12509"
                     /label="Protein Tat"
     promoter        8452..8546
                     /label="H1-2O2 promoter"
                     /note="doxycycline-inducible variant of the human H1 RNA
                     promoter (Henriksen et al., 2007)"
     misc_feature    8711..8828
                     /label="cPPT/CTS"
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
     promoter        8877..8880
                     /label="hPGK promoter"
                     /note="human phosphoglycerate kinase 1 promoter"
ORIGIN
        1 ttggggttgc gccttttcca aggcagccct gggtttgcgc agggacgcgg ctgctctggg
       61 cgtggttccg ggaaacgcag cggcgccgac cctgggtctc gcacattctt cacgtccgtt
      121 cgcagcgtca cccggatctt cgccgctacc cttgtgggcc ccccggcgac gcttcctgct
      181 ccgcccctaa gtcgggaagg ttccttgcgg ttcgcggcgt gccggacgtg acaaacggaa
      241 gccgcacgtc tcactagtac cctcgcagac ggacagcgcc agggagcaat ggcagcgcgc
      301 cgaccgcgat gggctgtggc caatagcggc tgctcagcag ggcgcgccga gagcagcggc
      361 cgggaagggg cggtgcggga ggcggggtgt ggggcggtag tgtgggccct gttcctgccc
      421 gcgcggtgtt ccgcattctg caagcctccg gagcgcacgt cggcagtcgg ctccctcgtt
      481 gaccgaatca ccgacctctc tccccagggg gatctgtgag tttggggacc cttgattgtt
      541 ctttcttttt cgctattgta aaattcatgt tatatggagg gggcaaagtt ttcagggtgt
      601 tgtttagaat gggaagatgt cccttgtatc accatggacc ctcatgataa ttttgtttct
      661 ttcactttct actctgttga caaccattgt ctcctcttat tttcttttca ttttctgtaa
      721 ctttttcgtt aaactttagc ttgcatttgt aacgaatttt taaattcact tttgtttatt
      781 tgtcagattg taagtacttt ctctaatcac ttttttttca aggcaatcag ggtatattat
      841 attgtacttc agcacagttt tagagaacaa ttgttataat taaatgataa ggtagaatat
      901 ttctgcatat aaattctggc tggcgtggaa atattcttat tggtagaaac aactacatcc
      961 tggtcatcat cctgcctttc tctttatggt tacaatgata tacactgttt gagatgagga
     1021 taaaatactc tgagtccaaa ccgggcccct ctgctaacca tgttcatgcc ttcttctttt
     1081 tcctacagct cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat
     1141 tgtaatacga ctcactatag ggcgaattga tatgtctaga ttagataaaa gtaaagtgat
     1201 taacagcgca ttagagctgc ttaatgaggt cggaatcgaa ggtttaacaa cccgtaaact
     1261 cgcccagaag ctaggtgtag agcagcctac attgtattgg catgtaaaaa ataagcgggc
     1321 tttgctcgac gccttagcca ttgagatgtt agataggcac catactcact tttgcccttt
     1381 agaaggggaa agctggcaag attttttacg taataacgct aaaagtttta gatgtgcttt
     1441 actaagtcat cgcgatggag caaaagtaca tttaggtaca cggcctacag aaaaacagta
     1501 tgaaactctc gaaaatcaat tagccttttt atgccaacaa ggtttttcac tagagaatgc
     1561 attatatgca ctcagcgctg tggggcattt tactttaggt tgcgtattgg aagatcaaga
     1621 gcatcaagtc gctaaagaag aaagggaaac acctactact gatagtatgc cgccattatt
     1681 acgacaagct atcgaattat ttgatcacca aggtgcagag ccagccttct tattcggcct
     1741 tgaattgatc atatgcggat tagaaaaaca acttaaatgt gaaagtgggt ccgcgtacag
     1801 cggctcccgg gagttctagg gatctgcccc tctccctccc ccccccctaa cgttactggc
     1861 cgaagccgct tggaataagg ccggtgtgcg tttgtctata tgttattttc caccatattg
     1921 ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg tcttcttgac gagcattcct
     1981 aggggtcttt cccctctcgc caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca
     2041 gttcctctgg aagcttcttg aagacaaaca acgtctgtag cgaccctttg caggcagcgg
     2101 aaccccccac ctggcgacag gtgcctctgc ggccaaaagc cacgtgtata agatacacct
     2161 gcaaaggcgg cacaacccca gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa
     2221 tggctctcct caagcgtatt caacaagggg ctgaaggatg cccagaaggt accccattgt
     2281 atgggatctg atctggggcc tcggtgcaca tgctttacat gtgtttagtc gaggttaaaa
     2341 aaacgtctag gccccccgaa ccacggggac gtggttttcc tttgaaaaac acgatgataa
     2401 ggatccaccg gagcttacca tgaccgagta caagcccacg gtgcgcctcg ccacccgcga
     2461 cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg
     2521 ccacaccgtc gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct
     2581 cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc
     2641 ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg
     2701 catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc
     2761 gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca
     2821 ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc
     2881 cggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct acgagcggct
     2941 cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac
     3001 ccgcaagccc ggtgcctgac gcccgcccca cgacccgcag cgcccgaccg aaaggagcgc
     3061 acgaccccat gcatcggtac ctttaagacc aatgacttac aaggcagctg tagatcttag
     3121 ccacttttta aaagaaaagg ggggactgga agggctaatt cactcccaac gaagacaaga
     3181 tctgcttttt gcttgtactg ggtctctctg gttagaccag atctgagcct gggagctctc
     3241 tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt
     3301 agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc
     3361 agtgtggaaa atctctagca gtagtagttc atgtcatctt attattcagt atttataact
     3421 tgcaaagaaa tgaatatcag agagtgagag gaacttgttt attgcagctt ataatggtta
     3481 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     3541 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggctctagc tatcccgccc
     3601 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     3661 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     3721 aagtagtgag gaggcttttt tggaggccta gggacgtacc caattcgccc tatagtgagt
     3781 cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg
     3841 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag
     3901 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc
     3961 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     4021 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     4081 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     4141 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     4201 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     4261 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     4321 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     4381 attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg
     4441 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     4501 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     4561 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     4621 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     4681 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     4741 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     4801 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     4861 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     4921 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     4981 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     5041 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     5101 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     5161 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     5221 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     5281 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     5341 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     5401 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     5461 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     5521 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     5581 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     5641 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     5701 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     5761 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     5821 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     5881 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     5941 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     6001 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     6061 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     6121 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     6181 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     6241 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     6301 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc
     6361 gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg
     6421 gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca
     6481 ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
     6541 tcacacagga aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag
     6601 ggaacaaaag ctggagctgc aagcttaatg tagtcttatg caatactctt gtagtcttgc
     6661 aacatggtaa cgatgagtta gcaacatgcc ttacaaggag agaaaaagca ccgtgcatgc
     6721 cgattggtgg aagtaaggtg gtacgatcgt gccttattag gaaggcaaca gacgggtctg
     6781 acatggattg gacgaaccac tgaattgccg cattgcagag atattgtatt taagtgccta
     6841 gctcgataca taaacgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     6901 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     6961 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     7021 tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg aaaccagagg
     7081 agctctctcg acgcaggact cggcttgctg aagcgcgcac ggcaagaggc gaggggcggc
     7141 gactggtgag tacgccaaaa attttgacta gcggaggcta gaaggagaga gatgggtgcg
     7201 agagcgtcag tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt cggttaaggc
     7261 cagggggaaa gaaaaaatat aaattaaaac atatagtatg ggcaagcagg gagctagaac
     7321 gattcgcagt taatcctggc ctgttagaaa catcagaagg ctgtagacaa atactgggac
     7381 agctacaacc atcccttcag acaggatcag aagaacttag atcattatat aatacagtag
     7441 caaccctcta ttgtgtgcat caaaggatag agataaaaga caccaaggaa gctttagaca
     7501 agatagagga agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc gctgatcttc
     7561 agacctggag gaggagatat gagggacaat tggagaagtg aattatataa atataaagta
     7621 gtaaaaattg aaccattagg agtagcaccc accaaggcaa agagaagagt ggtgcagaga
     7681 gaaaaaagag cagtgggaat aggagctttg ttccttgggt tcttgggagc agcaggaagc
     7741 actatgggcg cagcgtcaat gacgctgacg gtacaggcca gacaattatt gtctggtata
     7801 gtgcagcagc agaacaattt gctgagggct attgaggcgc aacagcatct gttgcaactc
     7861 acagtctggg gcatcaagca gctccaggca agaatcctgg ctgtggaaag atacctaaag
     7921 gatcaacagc tcctggggat ttggggttgc tctggaaaac tcatttgcac cactgctgtg
     7981 ccttggaatg ctagttggag taataaatct ctggaacaga tttggaatca cacgacctgg
     8041 atggagtggg acagagaaat taacaattac acaagcttaa tacactcctt aattgaagaa
     8101 tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg aattagataa atgggcaagt
     8161 ttgtggaatt ggtttaacat aacaaattgg ctgtggtata taaaattatt cataatgata
     8221 gtaggaggct tggtaggttt aagaatagtt tttgctgtac tttctatagt gaatagagtt
     8281 aggcagggat attcaccatt atcgtttcag acccacctcc caaccccgag gggacccgac
     8341 aggcccgaag gaatagaaga agaaggtgga gagagagaca gagacagatc cattcgatta
     8401 gtgaacggat ctcgacggta tcgatcacga gactagcctc gagcggccgc aatatttgca
     8461 tgtcgctatg tgttctggga aatcaccata aacgtgaaat ccctatcagt gatagagact
     8521 tataagttcc ctatcagtga tagagacacc ggttctgaac agcttcttgg gaagtagttt
     8581 aagagctaag ctggaaacag catagcaagt ttaaataagg ctagtccgtt atcaacttga
     8641 aaaagtggca ccgagtcggt gctttttttg aattctcgac ctcgagacaa atggcagtat
     8701 tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg gaaagaatag
     8761 tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt acaaaaattc
     8821 aaaattttcg ggtttattac agggacagca gagatccact ttggccgcgg ctcgaggggg
//
