LOCUS       sequence-239364-        6817 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of a non-targeting shRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6817)
  AUTHORS   Gaviraghi M, Vivori C, Pareja Sanchez Y, Invernizzi F, Cattaneo A, 
            Santoliquido BM, Frenquelli M, Segalla S, Bachi A, Doglioni C, 
            Pelechano V, Cittaro D, Tonon G
  TITLE     Tumor suppressor PNRC1 blocks rRNA maturation by recruiting the 
            decapping complex to the nucleolus.
  JOURNAL   EMBO J. 2018 Dec 3;37(23). pii: embj.201899179. doi: 
            10.15252/embj.201899179. Epub 2018 Oct 29.
  PUBMED    30373810
REFERENCE   2  (bases 1 to 6817)
  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: "EMBO J."; 
            date: "2018-12-3"; volume: "37(23). pii: embj.201899179. doi"; 
            pages: " 10.15252/embj.201899179. Epub 2018 Oct 29"
COMMENT     Sequence Label: pLKO.1 Blast NT shRNA
FEATURES             Location/Qualifiers
     source          1..6817
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    125..146
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(161..166,167..184,185..191)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    199..215
                     /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     204..226
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     223..239
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     223..239
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     258..278
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        260..278
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        306..532
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             533..713
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    760..885
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1378..1611
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1796..1840
                     /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        2138..2386
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2138..2158
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2309..2328
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    2483..2600
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2649..3159
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3078..3096
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3171..3569
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             3641..3874
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3946..4067
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3983..4002)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4037..4056
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4107..4242
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4262..4281)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4263..4281)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4291..4308)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4291..4307)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4300..4322)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4449..4904
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4536..4555)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4746..4767
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4930..5034
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5035..5103,5104..5895)
                     /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(5253..5272)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6066..6654
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6555..6574
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     join(6808..6817,1..8)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
       61 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg
      121 caattaatgt gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg
      181 ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc
      241 atgattacgc caagcgcgca attaaccctc actaaaggga acaaaagctg gagctgcaag
      301 cttaatgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga tgagttagca
      361 acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag taaggtggta
      421 cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac gaaccactga
      481 attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacataa acgggtctct
      541 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
      601 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
      661 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc
      721 ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg caggactcgg
      781 cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac gccaaaaatt
      841 ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat taagcggggg
      901 agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa
      961 ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa tcctggcctg
     1021 ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc ccttcagaca
     1081 ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg tgtgcatcaa
     1141 aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga gcaaaacaaa
     1201 agtaagacca ccgcacagca agcggccgct gatcttcaga cctggaggag gagatatgag
     1261 ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac cattaggagt
     1321 agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag tgggaatagg
     1381 agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag cgtcaatgac
     1441 gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga acaatttgct
     1501 gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca tcaagcagct
     1561 ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc tggggatttg
     1621 gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta gttggagtaa
     1681 taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca gagaaattaa
     1741 caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc aagaaaagaa
     1801 tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt ttaacataac
     1861 aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg taggtttaag
     1921 aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt caccattatc
     1981 gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa tagaagaaga
     2041 aggtggagag agagacagag acagatccat tcgattagtg aacggatctc gacggtatcg
     2101 atcacgagac tagcctcgag cggccgcccc cttcaccgag ggcctatttc ccatgattcc
     2161 ttcatatttg catatacgat acaaggctgt tagagagata attggaatta atttgactgt
     2221 aaacacaaag atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt
     2281 tgcagtttta aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta
     2341 tttcgatttc ttggctttat atatcttgtg gaaaggacga aacaccggca acaagatgaa
     2401 gagcaccaac tcgagttggt gctcttcatc ttgttgtttt tgaattctcg acctcgagac
     2461 aaatggcagt attcatccac aattttaaaa gaaaaggggg gattgggggg tacagtgcag
     2521 gggaaagaat agtagacata atagcaacag acatacaaac taaagaatta caaaaacaaa
     2581 ttacaaaaat tcaaaatttt cgggtttatt acagggacag cagagatcca ctttggccgc
     2641 ggctcgaggg ggttggggtt gcgccttttc caaggcagcc ctgggtttgc gcagggacgc
     2701 ggctgctctg ggcgtggttc cgggaaacgc agcggcgccg accctgggtc tcgcacattc
     2761 ttcacgtccg ttcgcagcgt cacccggatc ttcgccgcta cccttgtggg ccccccggcg
     2821 acgcttcctg ctccgcccct aagtcgggaa ggttccttgc ggttcgcggc gtgccggacg
     2881 tgacaaacgg aagccgcacg tctcactagt accctcgcag acggacagcg ccagggagca
     2941 atggcagcgc gccgaccgcg atgggctgtg gccaatagcg gctgctcagc agggcgcgcc
     3001 gagagcagcg gccgggaagg ggcggtgcgg gaggcggggt gtggggcggt agtgtgggcc
     3061 ctgttcctgc ccgcgcggtg ttccgcattc tgcaagcctc cggagcgcac gtcggcagtc
     3121 ggctccctcg ttgaccgaat caccgacctc tctccccagg gggatccacc atggccaagc
     3181 ctttgtctca agaagaatcc accctcattg aaagagcaac ggctacaatc aacagcatcc
     3241 ccatctctga agactacagc gtcgccagcg cagctctctc tagcgacggc cgcatcttca
     3301 ctggtgtcaa tgtatatcat tttactgggg gaccttgtgc agaactcgtg gtgctgggca
     3361 ctgctgctgc tgcggcagct ggcaacctga cttgtatcgt cgcgatcgga aatgagaaca
     3421 ggggcatctt gagcccctgc ggacggtgcc gacaggtgct tctcgatctg catcctggga
     3481 tcaaagccat agtgaaggac agtgatggac agccgacggc agttgggatt cgtgaattgc
     3541 tgccctctgg ttatgtgtgg gagggctaag gtacctttaa gaccaatgac ttacaaggca
     3601 gctgtagatc ttagccactt tttaaaagaa aaggggggac tggaagggct aattcactcc
     3661 caacgaagac aagatctgct ttttgcttgt actgggtctc tctggttaga ccagatctga
     3721 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
     3781 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
     3841 agaccctttt agtcagtgtg gaaaatctct agcagtagta gttcatgtca tcttattatt
     3901 cagtatttat aacttgcaaa gaaatgaata tcagagagtg agaggaactt gtttattgca
     3961 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     4021 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggctc
     4081 tagctatccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     4141 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
     4201 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctagggacg tacccaattc
     4261 gccctatagt gagtcgtatt acgcgcgctc actggccgtc gttttacaac gtcgtgactg
     4321 ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg
     4381 gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg
     4441 cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     4501 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     4561 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     4621 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg
     4681 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     4741 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt
     4801 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     4861 aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg
     4921 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     4981 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     5041 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     5101 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     5161 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     5221 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     5281 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     5341 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     5401 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     5461 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     5521 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     5581 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     5641 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     5701 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     5761 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     5821 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     5881 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     5941 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     6001 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     6061 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     6121 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     6181 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     6241 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     6301 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     6361 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     6421 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     6481 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     6541 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     6601 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     6661 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     6721 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     6781 gctcgccgca gccgaacgac cgagcgcagc gagtcag
//
