LOCUS       sequence-246021-        7084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  constitutive expression of a short-hairpin RNA targeting human PSMD2
            (RNAi positive control).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7084)
  AUTHORS   Chan EM, Shibue T, McFarland JM, Gaeta B, Ghandi M, Dumont N, 
            Gonzalez A, McPartlan JS, Li T, Zhang Y, Bin Liu J, Lazaro JB, Gu P,
            Piett CG, Apffel A, Ali SO, Deasy R, Keskula P, Ng RWS, Roberts EA, 
            Reznichenko E, Leung L, Alimova M, Schenone M, Islam M, Maruvka YE, 
            Liu Y, Roper J, Raghavan S, Giannakis M, Tseng YY, Nagel ZD, 
            D'Andrea A, Root DE, Boehm JS, Getz G, Chang S, Golub TR, Tsherniak 
            A, Vazquez F, Bass AJ
  TITLE     WRN helicase is a synthetic lethal target in microsatellite unstable
            cancers.
  JOURNAL   Nature. 2019 Apr 10. pii: 10.1038/s41586-019-1102-x. doi: 
            10.1038/s41586-019-1102-x.
  PUBMED    30971823
REFERENCE   2  (bases 1 to 7084)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature. 
            2019 Apr 10. pii: 10.1038/s41586-019-1102-x. doi: 
            10.1038/s41586-019-1102-x."
COMMENT     Sequence Label: pLKO.1-puro-shPSMD2
FEATURES             Location/Qualifiers
     source          1..7084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    88..109
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(124..129,130..147,148..154)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    162..178
                     /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     167..189
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     186..202
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     186..202
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     221..241
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        223..241
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        269..495
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             496..676
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    723..848
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1341..1574
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1759..1803
                     /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        2101..2349
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2101..2121
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2272..2291
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    2446..2563
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2612..3122
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3041..3059
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3144..3743
                     /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(3144..3163)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3640..3660
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3871..4104
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4176..4297
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4213..4232)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4267..4286
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4337..4472
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4492..4511)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4493..4511)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4521..4538)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4521..4537)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4530..4552)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4679..5134
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4766..4785)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4976..4997
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5160..5264
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5265..5333,5334..6125)
                     /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(5483..5502)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6296..6884
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6785..6804
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7038..7055
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg
       61 gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca
      121 ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
      181 tcacacagga aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag
      241 ggaacaaaag ctggagctgc aagcttaatg tagtcttatg caatactctt gtagtcttgc
      301 aacatggtaa cgatgagtta gcaacatgcc ttacaaggag agaaaaagca ccgtgcatgc
      361 cgattggtgg aagtaaggtg gtacgatcgt gccttattag gaaggcaaca gacgggtctg
      421 acatggattg gacgaaccac tgaattgccg cattgcagag atattgtatt taagtgccta
      481 gctcgataca taaacgggtc tctctggtta gaccagatct gagcctggga gctctctggc
      541 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
      601 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
      661 tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg aaaccagagg
      721 agctctctcg acgcaggact cggcttgctg aagcgcgcac ggcaagaggc gaggggcggc
      781 gactggtgag tacgccaaaa attttgacta gcggaggcta gaaggagaga gatgggtgcg
      841 agagcgtcag tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt cggttaaggc
      901 cagggggaaa gaaaaaatat aaattaaaac atatagtatg ggcaagcagg gagctagaac
      961 gattcgcagt taatcctggc ctgttagaaa catcagaagg ctgtagacaa atactgggac
     1021 agctacaacc atcccttcag acaggatcag aagaacttag atcattatat aatacagtag
     1081 caaccctcta ttgtgtgcat caaaggatag agataaaaga caccaaggaa gctttagaca
     1141 agatagagga agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc gctgatcttc
     1201 agacctggag gaggagatat gagggacaat tggagaagtg aattatataa atataaagta
     1261 gtaaaaattg aaccattagg agtagcaccc accaaggcaa agagaagagt ggtgcagaga
     1321 gaaaaaagag cagtgggaat aggagctttg ttccttgggt tcttgggagc agcaggaagc
     1381 actatgggcg cagcgtcaat gacgctgacg gtacaggcca gacaattatt gtctggtata
     1441 gtgcagcagc agaacaattt gctgagggct attgaggcgc aacagcatct gttgcaactc
     1501 acagtctggg gcatcaagca gctccaggca agaatcctgg ctgtggaaag atacctaaag
     1561 gatcaacagc tcctggggat ttggggttgc tctggaaaac tcatttgcac cactgctgtg
     1621 ccttggaatg ctagttggag taataaatct ctggaacaga tttggaatca cacgacctgg
     1681 atggagtggg acagagaaat taacaattac acaagcttaa tacactcctt aattgaagaa
     1741 tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg aattagataa atgggcaagt
     1801 ttgtggaatt ggtttaacat aacaaattgg ctgtggtata taaaattatt cataatgata
     1861 gtaggaggct tggtaggttt aagaatagtt tttgctgtac tttctatagt gaatagagtt
     1921 aggcagggat attcaccatt atcgtttcag acccacctcc caaccccgag gggacccgac
     1981 aggcccgaag gaatagaaga agaaggtgga gagagagaca gagacagatc cattcgatta
     2041 gtgaacggat ctcgacggta tcgatcacga gactagcctc gagcggccgc ccccttcacc
     2101 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
     2161 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
     2221 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
     2281 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
     2341 cgaaacaccg gcgccagtta gctcaatatc atctcgagat gatattgagc taactggcgt
     2401 ttttgaattc tcgacctcga gacaaatggc agtattcatc cacaatttta aaagaaaagg
     2461 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
     2521 aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
     2581 cagcagagat ccactttggc cgcggctcga gggggttggg gttgcgcctt ttccaaggca
     2641 gccctgggtt tgcgcaggga cgcggctgct ctgggcgtgg ttccgggaaa cgcagcggcg
     2701 ccgaccctgg gtctcgcaca ttcttcacgt ccgttcgcag cgtcacccgg atcttcgccg
     2761 ctacccttgt gggccccccg gcgacgcttc ctgctccgcc cctaagtcgg gaaggttcct
     2821 tgcggttcgc ggcgtgccgg acgtgacaaa cggaagccgc acgtctcact agtaccctcg
     2881 cagacggaca gcgccaggga gcaatggcag cgcgccgacc gcgatgggct gtggccaata
     2941 gcggctgctc agcagggcgc gccgagagca gcggccggga aggggcggtg cgggaggcgg
     3001 ggtgtggggc ggtagtgtgg gccctgttcc tgcccgcgcg gtgttccgca ttctgcaagc
     3061 ctccggagcg cacgtcggca gtcggctccc tcgttgaccg aatcaccgac ctctctcccc
     3121 agggggatcc accggagctt accatgaccg agtacaagcc cacggtgcgc ctcgccaccc
     3181 gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac taccccgcca
     3241 cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg caagaactct
     3301 tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg
     3361 tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc
     3421 cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg gaaggcctcc
     3481 tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg
     3541 accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag gcggccgagc
     3601 gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc ttctacgagc
     3661 ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc acctggtgca
     3721 tgacccgcaa gcccggtgcc tgacgcccgc cccacgaccc gcagcgcccg accgaaagga
     3781 gcgcacgacc ccatgcatcg gtacctttaa gaccaatgac ttacaaggca gctgtagatc
     3841 ttagccactt tttaaaagaa aaggggggac tggaagggct aattcactcc caacgaagac
     3901 aagatctgct ttttgcttgt actgggtctc tctggttaga ccagatctga gcctgggagc
     3961 tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc
     4021 aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt
     4081 agtcagtgtg gaaaatctct agcagtagta gttcatgtca tcttattatt cagtatttat
     4141 aacttgcaaa gaaatgaata tcagagagtg agaggaactt gtttattgca gcttataatg
     4201 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     4261 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggctc tagctatccc
     4321 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     4381 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt
     4441 ccagaagtag tgaggaggct tttttggagg cctagggacg tacccaattc gccctatagt
     4501 gagtcgtatt acgcgcgctc actggccgtc gttttacaac gtcgtgactg ggaaaaccct
     4561 ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc
     4621 gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatgggac
     4681 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     4741 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     4801 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     4861 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     4921 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     4981 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     5041 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     5101 gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc
     5161 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     5221 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     5281 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     5341 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     5401 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     5461 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     5521 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     5581 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     5641 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     5701 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     5761 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     5821 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     5881 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     5941 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     6001 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     6061 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     6121 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     6181 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     6241 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     6301 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     6361 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     6421 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     6481 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     6541 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     6601 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     6661 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     6721 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     6781 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     6841 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     6901 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     6961 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
     7021 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca
     7081 aacc
//
