LOCUS       V012234                10861 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  Exported.
ACCESSION   V012234
VERSION     V012234
KEYWORDS    pCLIP-Cas9-Nuclease-hCMV-Blast
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 10861)
  AUTHORS   transOMIC
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 10861)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..10861
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        27..404
                     /label="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     LTR             411..668
                     /label="5' LTR (truncated)"
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    715..840
                     /label="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    1333..1566
                     /label="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
     CDS             1751..1795
                     /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             1944..1985
                     /note="Protein Tat from Human immunodeficiency virus type 1
                     group M subtype B (isolate WMJ22). Accession#: P12509"
                     /label="Protein Tat"
     misc_feature    2057..2174
                     /label="cPPT/CTS"
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
     enhancer        2238..2541
                     /note="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2542..2745
                     /label="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     CDS             2905..7008
                     /label="Cas9"
                     /note="Cas9 (Csn1) endonuclease from the Streptococcus
                     pyogenes Type II CRISPR/Cas system"
     CDS             7009..7056
                     /codon_start=1
                     /product="bipartite nuclear localization signal from
                     nucleoplasmin"
                     /label="bipartite nuclear localization signal from nucl"
                     /note="nucleoplasmin NLS"
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7057..7080
                     /label="FLAG"
                     /note="FLAG(R) epitope tag, followed by an enterokinase
                     cleavage site"
     CDS             7090..7146
                     /label="P2A"
                     /note="2A peptide from porcine teschovirus-1 polyprotein"
     CDS             7147..7542
                     /label="BSD"
                     /note="blasticidin S deaminase"
     misc_feature    7570..8158
                     /label="WPRE"
                     /note="woodchuck hepatitis virus posttranscriptional
                     regulatory element"
     LTR             8363..8596
                     /label="3' LTR (Delta-U3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     promoter        8692..9021
                     /label="SV40 promoter"
                     /note="SV40 enhancer and early promoter"
     rep_origin      complement(9110..9695)
                     /direction=LEFT
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(9869..10726)
                     /label="AmpR"
                     /note="beta-lactamase"
     promoter        complement(10727..10831)
                     /label="AmpR promoter"
ORIGIN
        1 gacggatcgg gagatcctcg cgcgttgaca ttgattattg actagttatt aatagtaatc
       61 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
      121 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
      181 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
      241 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
      301 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
      361 tcctacttgg cagtacatct acgtattagt catcgctatt accagctagc gaggcgtggc
      421 ctgggcggga ctggggagtg gcgagccctc agatcctgca tataagcagc tgctttttgc
      481 ctgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg
      541 gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg
      601 tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat
      661 ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct
      721 cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg
      781 agtacgccaa aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc
      841 agtattaagc gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga
      901 aagaaaaaat ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca
      961 gttaatcctg gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa
     1021 ccatcccttc agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc
     1081 tattgtgtgc atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag
     1141 gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct tcagacctgg
     1201 aggaggagat atgagggaca attggagaag tgaattatat aaatataaag tagtaaaaat
     1261 tgaaccatta ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag
     1321 agcagtggga ataggagctt tgttccttgg gttcttggga gcagcaggaa gcactatggg
     1381 cgcagcgtca atgacgctga cggtacaggc cagacaatta ttgtctggta tagtgcagca
     1441 gcagaacaat ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac tcacagtctg
     1501 gggcatcaag cagctccagg caagaatcct ggctgtggaa agatacctaa aggatcaaca
     1561 gctcctgggg atttggggtt gctctggaaa actcatttgc accactgctg tgccttggaa
     1621 tgctagttgg agtaataaat ctctggaaca gatttggaat cacacgacct ggatggagtg
     1681 ggacagagaa attaacaatt acacaagctt aatacactcc ttaattgaag aatcgcaaaa
     1741 ccagcaagaa aagaatgaac aagaattatt ggaattagat aaatgggcaa gtttgtggaa
     1801 ttggtttaac ataacaaatt ggctgtggta tataaaatta ttcataatga tagtaggagg
     1861 cttggtaggt ttaagaatag tttttgctgt actttctata gtgaatagag ttaggcaggg
     1921 atattcacca ttatcgtttc agacccacct cccaaccccg aggggacccg acaggcccga
     1981 aggaatagaa gaagaaggtg gagagagaga cagagacaga tccattcgat tagtgaacgg
     2041 atctcgacgg tatcgatttt aaaagaaaag gggggattgg ggggtacagt gcaggggaaa
     2101 gaatagtaga cataatagca acagacatac aaactaaaga actacaaaaa caaattacaa
     2161 aaattcaaaa ttttcgggtt tattacaggg acagcagaga tccagtttgg aattcatcga
     2221 taagcttggg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
     2281 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
     2341 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
     2401 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
     2461 tggcattatg cccagtacat gaccttacgg gactttccta cttggcagta catctacgta
     2521 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acaccaatgg gcgtggatag
     2581 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
     2641 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
     2701 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctcgttt agtgaaccgt
     2761 cagatcgcct ggagacgcca tccacgctgt tttgacctcc atagaagaca ccgactctag
     2821 ctagaggatc cactagtcca gtgtggtgga attctgcaga tatccagcac agtggcggcc
     2881 gctcgagtct agagcgctgc caccatggac aagaagtaca gcatcggcct ggacatcggc
     2941 accaactctg tgggctgggc cgtgatcacc gacgagtaca aggtgcccag caagaaattc
     3001 aaggtgctgg gcaacaccga ccggcacagc atcaagaaga acctgatcgg agccctgctg
     3061 ttcgacagcg gcgaaacagc cgaggccacc cggctgaaga gaaccgccag aagaagatac
     3121 accagacgga agaaccggat ctgctatctg caagagatct tcagcaacga gatggccaag
     3181 gtggacgaca gcttcttcca cagactggaa gagtccttcc tggtggaaga ggataagaag
     3241 cacgagcggc accccatctt cggcaacatc gtggacgagg tggcctacca cgagaagtac
     3301 cccaccatct accacctgag aaagaaactg gtggacagca ccgacaaggc cgacctgcgg
     3361 ctgatctatc tggccctggc ccacatgatc aagttccggg gccacttcct gatcgagggc
     3421 gacctgaacc ccgacaacag cgacgtggac aagctgttca tccagctggt gcagacctac
     3481 aaccagctgt tcgaggaaaa ccccatcaac gccagcggcg tggacgccaa ggccatcctg
     3541 tctgccagac tgagcaagag cagacggctg gaaaatctga tcgcccagct gcccggcgag
     3601 aagaagaatg gcctgttcgg aaacctgatt gccctgagcc tgggcctgac ccccaacttc
     3661 aagagcaact tcgacctggc cgaggatgcc aaactgcagc tgagcaagga cacctacgac
     3721 gacgacctgg acaacctgct ggcccagatc ggcgaccagt acgccgacct gtttctggcc
     3781 gccaagaacc tgtccgacgc catcctgctg agcgacatcc tgagagtgaa caccgagatc
     3841 accaaggccc ccctgagcgc ctctatgatc aagagatacg acgagcacca ccaggacctg
     3901 accctgctga aagctctcgt gcggcagcag ctgcctgaga agtacaaaga gattttcttc
     3961 gaccagagca agaacggcta cgccggctac attgacggcg gagccagcca ggaagagttc
     4021 tacaagttca tcaagcccat cctggaaaag atggacggca ccgaggaact gctcgtgaag
     4081 ctgaacagag aggacctgct gcggaagcag cggaccttcg acaacggcag catcccccac
     4141 cagatccacc tgggagagct gcacgccatt ctgcggcggc aggaagattt ttacccattc
     4201 ctgaaggaca accgggaaaa gatcgagaag atcctgacct tccgcatccc ctactacgtg
     4261 ggccctctgg ccaggggaaa cagcagattc gcctggatga ccagaaagag cgaggaaacc
     4321 atcaccccct ggaacttcga ggaagtggtg gacaagggcg cttccgccca gagcttcatc
     4381 gagcggatga ccaacttcga taagaacctg cccaacgaga aggtgctgcc caagcacagc
     4441 ctgctgtacg agtacttcac cgtgtataac gagctgacca aagtgaaata cgtgaccgag
     4501 ggaatgagaa agcccgcctt cctgagcggc gagcagaaaa aggccatcgt ggacctgctg
     4561 ttcaagacca accggaaagt gaccgtgaag cagctgaaag aggactactt caagaaaatc
     4621 gagtgcttcg actccgtgga aatctccggc gtggaagatc ggttcaacgc ctccctgggc
     4681 acataccacg atctgctgaa aattatcaag gacaaggact tcctggacaa tgaggaaaac
     4741 gaggacattc tggaagatat cgtgctgacc ctgacactgt ttgaggacag agagatgatc
     4801 gaggaacggc tgaaaaccta tgcccacctg ttcgacgaca aagtgatgaa gcagctgaag
     4861 cggcggagat acaccggctg gggcaggctg agccggaagc tgatcaacgg catccgggac
     4921 aagcagtccg gcaagacaat cctggatttc ctgaagtccg acggcttcgc caacagaaac
     4981 ttcatgcagc tgatccacga cgacagcctg acctttaaag aggacatcca gaaagcccag
     5041 gtgtccggcc agggcgatag cctgcacgag cacattgcca atctggccgg cagccccgcc
     5101 attaagaagg gcatcctgca gacagtgaag gtggtggacg agctcgtgaa agtgatgggc
     5161 cggcacaagc ccgagaacat cgtgatcgaa atggccagag agaaccagac cacccagaag
     5221 ggacagaaga acagccgcga gagaatgaag cggatcgaag agggcatcaa agagctgggc
     5281 agccagatcc tgaaagaaca ccccgtggaa aacacccagc tgcagaacga gaagctgtac
     5341 ctgtactacc tgcagaatgg gcgggatatg tacgtggacc aggaactgga catcaaccgg
     5401 ctgtccgact acgatgtgga ccatatcgtg cctcagagct ttctgaagga cgactccatc
     5461 gacaacaagg tgctgaccag aagcgacaag aaccggggca agagcgacaa cgtgccctcc
     5521 gaagaggtcg tgaagaagat gaagaactac tggcggcagc tgctgaacgc caagctgatt
     5581 acccagagaa agttcgacaa tctgaccaag gccgagagag gcggcctgag cgaactggat
     5641 aaggccggct tcatcaagag acagctggtg gaaacccggc agatcacaaa gcacgtggca
     5701 cagatcctgg actcccggat gaacactaag tacgacgaga atgacaagct gatccgggaa
     5761 gtgaaagtga tcaccctgaa gtccaagctg gtgtccgatt tccggaagga tttccagttt
     5821 tacaaagtgc gcgagatcaa caactaccac cacgcccacg acgcctacct gaacgccgtc
     5881 gtgggaaccg ccctgatcaa aaagtaccct aagctggaaa gcgagttcgt gtacggcgac
     5941 tacaaggtgt acgacgtgcg gaagatgatc gccaagagcg agcaggaaat cggcaaggct
     6001 accgccaagt acttcttcta cagcaacatc atgaactttt tcaagaccga gattaccctg
     6061 gccaacggcg agatccggaa gcggcctctg atcgagacaa acggcgaaac cggggagatc
     6121 gtgtgggata agggccggga ttttgccacc gtgcggaaag tgctgagcat gccccaagtg
     6181 aatatcgtga aaaagaccga ggtgcagaca ggcggcttca gcaaagagtc tatcctgccc
     6241 aagaggaaca gcgataagct gatcgccaga aagaaggact gggaccctaa gaagtacggc
     6301 ggcttcgaca gccccaccgt ggcctattct gtgctggtgg tggccaaagt ggaaaagggc
     6361 aagtccaaga aactgaagag tgtgaaagag ctactgggga tcaccatcat ggaaagaagc
     6421 agcttcgaga agaatcccat cgactttctg gaagccaagg gctacaaaga agtgaaaaag
     6481 gacctgatca tcaagctgcc taagtactcc ctgttcgagc tggaaaacgg ccggaagaga
     6541 atgctggcct ctgccggcga actgcagaag ggaaacgaac tggccctgcc ctccaaatat
     6601 gtgaacttcc tgtacctggc cagccactat gagaagctga agggctcccc cgaggataat
     6661 gagcagaaac agctgtttgt ggaacagcac aagcactacc tggacgagat catcgagcag
     6721 atcagcgagt tctccaagag agtgatcctg gccgacgcta atctggacaa agtgctgtcc
     6781 gcctacaaca agcaccggga taagcccatc agagagcagg ccgagaatat catccacctg
     6841 tttaccctga ccaatctggg agcccctgcc gccttcaagt actttgacac caccatcgac
     6901 cggaagaggt acaccagcac caaagaggtg ctggacgcca ccctgatcca ccagagcatc
     6961 accggcctgt acgagacacg gatcgacctg tctcagctgg gaggcgacaa gcgacctgcc
     7021 gccacaaaga aggctggaca ggctaagaag aagaaagatt acaaagacga tgacgataag
     7081 ggatccggcg caacaaactt ctctctgctg aaacaagccg gagatgtcga agagaatcct
     7141 ggaccgatgg ccaagccttt gtctcaagaa gaatccaccc tcattgaaag agcaacggct
     7201 acaatcaaca gcatccccat ctctgaagac tacagcgtcg ccagcgcagc tctctctagc
     7261 gacggccgca tcttcactgg tgtcaatgta tatcatttta ctgggggacc ttgtgcagaa
     7321 ctcgtggtgc tgggcactgc tgctgctgcg gcagctggca acctgacttg tatcgtcgcg
     7381 atcggaaatg agaacagggg catcttgagc ccctgcggac ggtgccgaca ggtgcttctc
     7441 gatctgcatc ctgggatcaa agccatagtg aaggacagtg atggacagcc gacggcagtt
     7501 gggattcgtg aattgctgcc ctctggttat gtgtgggagg gctaagaatt cgatatcaag
     7561 cttatcggta atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac
     7621 tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt
     7681 gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat
     7741 gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca
     7801 acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc
     7861 cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg
     7921 gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct
     7981 tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct
     8041 tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt
     8101 ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcat
     8161 cgataccgtc gacctcgaga cctagaaaaa catggagcaa tcacaagtag caatacagca
     8221 gctaccaatg ctgattgtgc ctggctagaa gcacaagagg aggaggaggt gggttttcca
     8281 gtcacacctc aggtaccttt aagaccaatg acttacaagg cagctgtaga tcttagccac
     8341 tttttaaaag aaaagagggg actggaaggg ctaattcact cccaacgaag acaagatctg
     8401 ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     8461 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     8521 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     8581 tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt ataacttgca
     8641 aagaaatgaa tatcagagag tgagaggcct tgacattgtt taaactggaa tgtgtgtcag
     8701 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     8761 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     8821 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     8881 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     8941 gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt
     9001 ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccattac atgtgagcaa
     9061 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     9121 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     9181 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     9241 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     9301 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     9361 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     9421 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     9481 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     9541 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     9601 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtttt tttgtttgca
     9661 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     9721 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     9781 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     9841 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     9901 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     9961 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
    10021 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
    10081 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
    10141 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
    10201 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
    10261 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
    10321 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
    10381 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
    10441 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
    10501 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
    10561 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
    10621 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
    10681 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
    10741 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
    10801 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
    10861 t
//
