LOCUS       Exported                8901 bp ds-DNA     circular SYN 27-OCT-2023
DEFINITION  3rd generation transfer plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    pLKO.1 - TRC cloning vector
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8901)
  AUTHORS   Moffat J, Grueneberg DA, Yang X, Kim SY, Kloepfer AM, Hinkle G, 
            Piqani B, Eisenhaure TM, Luo B, Grenier JK, Carpenter AE, Foo SY, 
            Stewart SA, Stockwell BR, Hacohen N, Hahn WC, Lander ES, Sabatini 
            DM, Root DE
  TITLE     A lentiviral RNAi library for human and mouse genes applied to an 
            arrayed viral high-content screen.
  JOURNAL   Cell. 2006 Mar 24. 124(6):1283-98.
  PUBMED    16564017
REFERENCE   2  (bases 1 to 8901)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 27, 2023 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8901
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     77..94
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    211..232
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        247..277
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    285..301
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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     290..312
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     309..325
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     309..325
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     344..364
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        346..364
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        392..618
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             619..799
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    846..971
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1464..1697
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1882..1926
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     promoter        2224..2464
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     2224..2244
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2395..2414
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    4386..4503
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        4552..5062
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     4981..4999
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             5084..5683
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5084..5103)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5580..5600
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             5811..6044
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    6116..6237
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6153..6172)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6207..6226
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      6277..6412
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6339..6358
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(6432..6451)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6433..6451)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6461..6478)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6461..6477)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6470..6492)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6619..7074
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6706..6725)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6916..6937
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        7100..7204
                     /gene="bla"
                     /label=AmpR promoter
     CDS             7205..8065
                     /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(7423..7442)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8236..8824
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8725..8744
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
       61 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
      121 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
      181 ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc
      241 ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca
      301 atttcacaca ggaaacagct atgaccatga ttacgccaag cgcgcaatta accctcacta
      361 aagggaacaa aagctggagc tgcaagctta atgtagtctt atgcaatact cttgtagtct
      421 tgcaacatgg taacgatgag ttagcaacat gccttacaag gagagaaaaa gcaccgtgca
      481 tgccgattgg tggaagtaag gtggtacgat cgtgccttat taggaaggca acagacgggt
      541 ctgacatgga ttggacgaac cactgaattg ccgcattgca gagatattgt atttaagtgc
      601 ctagctcgat acataaacgg gtctctctgg ttagaccaga tctgagcctg ggagctctct
      661 ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta
      721 gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca
      781 gtgtggaaaa tctctagcag tggcgcccga acagggactt gaaagcgaaa gggaaaccag
      841 aggagctctc tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc
      901 ggcgactggt gagtacgcca aaaattttga ctagcggagg ctagaaggag agagatgggt
      961 gcgagagcgt cagtattaag cgggggagaa ttagatcgcg atgggaaaaa attcggttaa
     1021 ggccaggggg aaagaaaaaa tataaattaa aacatatagt atgggcaagc agggagctag
     1081 aacgattcgc agttaatcct ggcctgttag aaacatcaga aggctgtaga caaatactgg
     1141 gacagctaca accatccctt cagacaggat cagaagaact tagatcatta tataatacag
     1201 tagcaaccct ctattgtgtg catcaaagga tagagataaa agacaccaag gaagctttag
     1261 acaagataga ggaagagcaa aacaaaagta agaccaccgc acagcaagcg gccgctgatc
     1321 ttcagacctg gaggaggaga tatgagggac aattggagaa gtgaattata taaatataaa
     1381 gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg caaagagaag agtggtgcag
     1441 agagaaaaaa gagcagtggg aataggagct ttgttccttg ggttcttggg agcagcagga
     1501 agcactatgg gcgcagcgtc aatgacgctg acggtacagg ccagacaatt attgtctggt
     1561 atagtgcagc agcagaacaa tttgctgagg gctattgagg cgcaacagca tctgttgcaa
     1621 ctcacagtct ggggcatcaa gcagctccag gcaagaatcc tggctgtgga aagataccta
     1681 aaggatcaac agctcctggg gatttggggt tgctctggaa aactcatttg caccactgct
     1741 gtgccttgga atgctagttg gagtaataaa tctctggaac agatttggaa tcacacgacc
     1801 tggatggagt gggacagaga aattaacaat tacacaagct taatacactc cttaattgaa
     1861 gaatcgcaaa accagcaaga aaagaatgaa caagaattat tggaattaga taaatgggca
     1921 agtttgtgga attggtttaa cataacaaat tggctgtggt atataaaatt attcataatg
     1981 atagtaggag gcttggtagg tttaagaata gtttttgctg tactttctat agtgaataga
     2041 gttaggcagg gatattcacc attatcgttt cagacccacc tcccaacccc gaggggaccc
     2101 gacaggcccg aaggaataga agaagaaggt ggagagagag acagagacag atccattcga
     2161 ttagtgaacg gatctcgacg gtatcgatca cgagactagc ctcgagcggc cgcccccttc
     2221 accgagggcc tatttcccat gattccttca tatttgcata tacgatacaa ggctgttaga
     2281 gagataattg gaattaattt gactgtaaac acaaagatat tagtacaaaa tacgtgacgt
     2341 agaaagtaat aatttcttgg gtagtttgca gttttaaaat tatgttttaa aatggactat
     2401 catatgctta ccgtaacttg aaagtatttc gatttcttgg ctttatatat cttgtggaaa
     2461 ggacgaaaca ccggtgtgtt gtaaatgagc acacaaaata cacatgctaa aatattatat
     2521 tctatgacct ttataaaatc aaccaaaatc ttctttttaa taactttagt atcaataatt
     2581 agaattttta tgttcctttt tgcaaacttt taataaaaat gagcaaaata aaaaaacgct
     2641 agttttagta actcgcgttg ttttcttcac ctttaataat agctactcca ccacttgttc
     2701 ctaagcggtc agctcctgct tcaatcattt tttgagcatc ttcaaatgtt ctaactccac
     2761 cagctgcttt aactaaagca ttgtctttaa caactgactt cattagttta acatcttcaa
     2821 atgttgcacc tgattttgaa aatcctgttg atgttttaac aaattctaat ccagcttcaa
     2881 cagctatttc acaagctttc atgatttctt cttttgttaa taaacaattt tccataatac
     2941 atttaacaac atgtgatcca gctgcttttt ttacagcttt catgtcttct aaaactaatt
     3001 cataattttt gtcttttaat gcaccaatat ttaataccat atcaatttct gttgcaccat
     3061 ctttaattgc ttcagaaact tcgaatgctt ttgtagctgt tgtgcatgca cctagaggaa
     3121 aacctacaac atttgttatt cctacatttg tgccttttaa taattcttta caatagcttg
     3181 ttcaatatga attaacacaa actgttgcaa aatcaaattc aattgcttca tcacataatt
     3241 gtttaatttc agctttcgta gcatcttgtt ttaataatgt gtgatctata tatttgttta
     3301 gtttcatttt ttctcctata tattcatttt taattttaat tctttaataa tttcgtctac
     3361 tttaacttta gcgttttgaa cagattcacc aacacctata aaataaattt ttagtttagg
     3421 ttcagttcca cttgggcgaa cagcaaatca tgacttatct tctaaataaa attttagtaa
     3481 gtcttgtcct ggcatattat acattccatc gatgtagtct tcaacattaa caactttaag
     3541 tccagcaatt tgagttaagg gtgttgctct caatgatttc attaatggtt caatttttaa
     3601 tttcttttct tctggtttaa aattcaagtt taaagtgaaa gtgtaatatg cacccatttc
     3661 tttaaataaa tcttctaaat agtctactaa tgttttattt tgttttttat aaaatcaagc
     3721 agcctctgct attaatatag aagcttgtat tccatcttta tctctagctg agtcatcaat
     3781 tacatatcca taactttctt cataagcaaa aacaaaattt aatccgttat cttcttcttt
     3841 agcaatttct ctacccattc atttaaatcc agttaaagtt tttacaatat taactccata
     3901 tttttcatga gcgattctat cacccaaatc acttgttaca aaacttgaat atagagccgg
     3961 attttttgga atgctattta agcgttttag atttgataat tttcaatcaa ttaaaattgg
     4021 tcctgtttga tttccatcta atcttacaaa atgaccatca tgttttattg ccattccaaa
     4081 tctgtcagca tctgggtcat tcataataat aatatctgca tcatgtttaa taccatattc
     4141 aagcggtatt tttcatgcag gatcaaattc tggatttgga tttacaacat ttttaaatgt
     4201 ttcatcttca aatgcatgct cttcaacctc aataacgtta tatcctgatt cacgtaatat
     4261 ttttggggta aatttagttc ctgttccatt aactgcgcta aaaataattt ttaaatcttt
     4321 tttagcttct tgctcttttt tgtagaattc tcgacctcga gacaaatggc agtattcatc
     4381 cacaatttta aaagaaaagg ggggattggg gggtacagtg caggggaaag aatagtagac
     4441 ataatagcaa cagacataca aactaaagaa ttacaaaaac aaattacaaa aattcaaaat
     4501 tttcgggttt attacaggga cagcagagat ccactttggc cgcggctcga gggggttggg
     4561 gttgcgcctt ttccaaggca gccctgggtt tgcgcaggga cgcggctgct ctgggcgtgg
     4621 ttccgggaaa cgcagcggcg ccgaccctgg gtctcgcaca ttcttcacgt ccgttcgcag
     4681 cgtcacccgg atcttcgccg ctacccttgt gggccccccg gcgacgcttc ctgctccgcc
     4741 cctaagtcgg gaaggttcct tgcggttcgc ggcgtgccgg acgtgacaaa cggaagccgc
     4801 acgtctcact agtaccctcg cagacggaca gcgccaggga gcaatggcag cgcgccgacc
     4861 gcgatgggct gtggccaata gcggctgctc agcagggcgc gccgagagca gcggccggga
     4921 aggggcggtg cgggaggcgg ggtgtggggc ggtagtgtgg gccctgttcc tgcccgcgcg
     4981 gtgttccgca ttctgcaagc ctccggagcg cacgtcggca gtcggctccc tcgttgaccg
     5041 aatcaccgac ctctctcccc agggggatcc accggagctt accatgaccg agtacaagcc
     5101 cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc
     5161 gttcgccgac taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt
     5221 caccgagctg caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt
     5281 cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc
     5341 ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca
     5401 gcaacagatg gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc
     5461 caccgtcggc gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc
     5521 cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg
     5581 caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga
     5641 aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgacgcccgc cccacgaccc
     5701 gcagcgcccg accgaaagga gcgcacgacc ccatgcatcg gtacctttaa gaccaatgac
     5761 ttacaaggca gctgtagatc ttagccactt tttaaaagaa aaggggggac tggaagggct
     5821 aattcactcc caacgaagac aagatctgct ttttgcttgt actgggtctc tctggttaga
     5881 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata
     5941 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta
     6001 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtagta gttcatgtca
     6061 tcttattatt cagtatttat aacttgcaaa gaaatgaata tcagagagtg agaggaactt
     6121 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     6181 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     6241 tgtctggctc tagctatccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     6301 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     6361 gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctagggacg
     6421 tacccaattc gccctatagt gagtcgtatt acgcgcgctc actggccgtc gttttacaac
     6481 gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt
     6541 tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca
     6601 gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     6661 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     6721 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     6781 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     6841 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     6901 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     6961 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     7021 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg
     7081 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     7141 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     7201 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     7261 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     7321 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     7381 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     7441 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     7501 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     7561 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     7621 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     7681 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     7741 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     7801 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     7861 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     7921 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     7981 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     8041 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     8101 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     8161 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     8221 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     8281 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     8341 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     8401 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     8461 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     8521 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     8581 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     8641 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     8701 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     8761 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     8821 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     8881 catgttcttt cctgcgttat c
//
