LOCUS       sequence-472027-        8421 bp    DNA     circular SYN 07-MAY-2025
DEFINITION  shRNA against human USP7.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8421)
  TITLE     Manel lab plasmids 2025
REFERENCE   2  (bases 1 to 8421)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 7, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pLKO.1-puro-USP7-sh1
FEATURES             Location/Qualifiers
     source          1..8421
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     426..444
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             529..1128
                     /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(529..548)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1025..1045
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             1256..1489
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1561..1682
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1598..1617)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1652..1671
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      1722..1857
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1877..1896)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1878..1896)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1906..1923)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1906..1922)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1915..1937)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2064..2519
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2151..2170)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2361..2382
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2545..2649
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2650..2718,2719..3510)
                     /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(2868..2887)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3681..4269
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4170..4189
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4423..4440
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4557..4578
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4593..4598,4599..4616,4617..4623)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4631..4647
                     /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     4636..4658
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4655..4671
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4655..4671
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4690..4710
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        4692..4710
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6076..6302
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6303..6483
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6530..6655
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7148..7381
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7566..7610
                     /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        7908..8156
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     7908..7928
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     8079..8098
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    8252..8369
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        join(8418..8421,1..507)
                     /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 gatccaccgg agcttaccat gaccgagtac
      541 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
      601 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
      661 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
      721 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
      781 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
      841 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
      901 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
      961 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg
     1021 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
     1081 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgacg cccgccccac
     1141 gacccgcagc gcccgaccga aaggagcgca cgaccccatg catcggtacc tttaagacca
     1201 atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa
     1261 gggctaattc actcccaacg aagacaagat ctgctttttg cttgtactgg gtctctctgg
     1321 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
     1381 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
     1441 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag tagtagttca
     1501 tgtcatctta ttattcagta tttataactt gcaaagaaat gaatatcaga gagtgagagg
     1561 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     1621 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     1681 tatcatgtct ggctctagct atcccgcccc taactccgcc catcccgccc ctaactccgc
     1741 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     1801 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     1861 ggacgtaccc aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt
     1921 acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc
     1981 ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt
     2041 gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     2101 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     2161 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     2221 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     2281 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     2341 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     2401 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     2461 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
     2521 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     2581 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     2641 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     2701 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     2761 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     2821 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     2881 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     2941 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     3001 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     3061 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     3121 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     3181 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     3241 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     3301 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     3361 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     3421 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     3481 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     3541 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     3601 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     3661 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     3721 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     3781 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     3841 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     3901 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     3961 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     4021 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     4081 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     4141 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     4201 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     4261 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     4321 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     4381 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     4441 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
     4501 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
     4561 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
     4621 ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac
     4681 gccaagcgcg caattaaccc tcactaaagg gaacaaaagc tggagctgca agcttaatgc
     4741 tgatgaatcc cctaatgatt tttatcaaaa tcattaagtt aaggtagata cacatcttgt
     4801 catatgatca aatggtttcg ccaaaaatca ataatcagac aacaaaatgt gcgaactcga
     4861 tattttacac gactctcttt accaattctg ccccgaatta cacttaaaac gactcaacag
     4921 cttaacgttg gcttgccacg ccttacttga ctgtaaaact ctcactctta ccgaacttgg
     4981 ccgtaacctg ccaaccaaag cgagaacaaa acataacatc aaacgaatcg accgattgtt
     5041 aggtaatcgt cacctccaca aagagcgact cgctgtatac cgttggcatg ctagctttat
     5101 ctgttcgggc aatacgatgc ccattgtact tgttgactgg tctgatatcc gtgagcaaaa
     5161 acggcttatg gtattgcgag cttcagtcgc actacacggt cgttctgtta ctctttatga
     5221 gaaagcgttc ccgctttcag agcaatgttc aaagaaagct catgaccaat ttctagccga
     5281 ccttgcgagc attctaccga gtaacaccac accgctcatt gtcagtgatg ctggctttaa
     5341 agtgccatgg tataaatccg ttgagaagct gggttggtac tggttaagtc gagtaagagg
     5401 aaaagtacaa tatgcagacc taggagcgga aaactggaaa cctatcagca acttacatga
     5461 tatgtcatct agtcactcaa agactttagg ctataagagg ctgactaaaa gcaacccaat
     5521 ctcatgccaa attctattgt ataaatctcg ctctaaaggc cgaaaaaatc agcgctcgac
     5581 acggactcat tgtcaccacc cgtcacctaa aatctactca gcgtcggcaa aggagccatg
     5641 gattctagca actaacttac ctgttgaaat tcgaacaccc aaacaacttg ttaatatcta
     5701 ttcgaagcga atgcagattg aagaaacctt ccgagacttg aaaagtcctg cctacggact
     5761 aggcctacgc catagccgaa cgagcagctc agagcgtttt gatatcatgc tgctaatcgc
     5821 cctgatgctt caactaacat gttggcttgc gggcgttcat gctcagaaac aaggttggga
     5881 caagcacttc caggctaaca cagtcagaaa tcgaaacgta ctctcaacag ttcgcttagg
     5941 catggaagtt ttgcggcatt ctggctacac aataacaagg gaagactcac tcgtggctgc
     6001 aaccctgctt actcaaaatc tattcacaca tggttacgtt ttggggaaat tatgagggga
     6061 tctctcagag cttaatgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga
     6121 tgagttagca acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag
     6181 taaggtggta cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac
     6241 gaaccactga attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacataa
     6301 acgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc
     6361 cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt
     6421 tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta
     6481 gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg
     6541 caggactcgg cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac
     6601 gccaaaaatt ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat
     6661 taagcggggg agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa
     6721 aaaatataaa ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa
     6781 tcctggcctg ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc
     6841 ccttcagaca ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg
     6901 tgtgcatcaa aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga
     6961 gcaaaacaaa agtaagacca ccgcacagca agcggccgct gatcttcaga cctggaggag
     7021 gagatatgag ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac
     7081 cattaggagt agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag
     7141 tgggaatagg agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag
     7201 cgtcaatgac gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga
     7261 acaatttgct gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca
     7321 tcaagcagct ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc
     7381 tggggatttg gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta
     7441 gttggagtaa taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca
     7501 gagaaattaa caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc
     7561 aagaaaagaa tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt
     7621 ttaacataac aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg
     7681 taggtttaag aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt
     7741 caccattatc gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa
     7801 tagaagaaga aggtggagag agagacagag acagatccat tcgattagtg aacggatctc
     7861 gacggtatcg atcacgagac tagcctcgag cggccgcccc cttcaccgag ggcctatttc
     7921 ccatgattcc ttcatatttg catatacgat acaaggctgt tagagagata attggaatta
     7981 atttgactgt aaacacaaag atattagtac aaaatacgtg acgtagaaag taataatttc
     8041 ttgggtagtt tgcagtttta aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa
     8101 cttgaaagta tttcgatttc ttggctttat atatcttgtg gaaaggacga aacaccggcc
     8161 agctaagtat caaaggaaac tcgagtttcc tttgatactt agctggtttt taattctcga
     8221 cctcgagaca aatggcagta ttcatccaca attttaaaag aaaagggggg attggggggt
     8281 acagtgcagg ggaaagaata gtagacataa tagcaacaga catacaaact aaagaattac
     8341 aaaaacaaat tacaaaaatt caaaattttc gggtttatta cagggacagc agagatccac
     8401 tttggccgcg gctcgagggg g
//
