LOCUS       sequence-221597-        7084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector for expression of shRNA sequence targeting mouse 
            Nsd1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7084)
  AUTHORS   Streubel G, Watson A, Jammula SG, Scelfo A, Fitzpatrick DJ, Oliviero
            G, McCole R, Conway E, Glancy E, Negri GL, Dillon E, Wynne K, Pasini
            D, Krogan NJ, Bracken AP, Cagney G
  TITLE     The H3K36me2 Methyltransferase Nsd1 Demarcates PRC2-Mediated 
            H3K27me2 and H3K27me3 Domains in Embryonic Stem Cells.
  JOURNAL   Mol Cell. 2018 Apr 19;70(2):371-379.e5. doi: 
            10.1016/j.molcel.2018.02.027. Epub 2018 Mar 29.
  PUBMED    29606589
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"; doi: 
            "10.1016/j.molcel.2018.02.027"; journalName: "Mol Cell"; date: 
            "2018-04-19- 19"; volume: "70"; issue: "2"; pages: "371-379.e5"
COMMENT     Sequence Label: pLKO.TRC1.shmNsd1.1, puro
FEATURES             Location/Qualifiers
     source          1..7084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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        join(247..252,253..270,271..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..2472
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     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    2569..2686
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2735..3245
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3164..3182
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3267..3866
                     /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(3267..3286)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3763..3783
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3994..4227
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4299..4420
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4336..4355)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4390..4409
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4460..4595
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4615..4634)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4616..4634)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4644..4661)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4644..4660)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4653..4675)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4802..5257
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4889..4908)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5099..5120
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5283..5387
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5388..5456,5457..6248)
                     /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(5606..5625)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6419..7007
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6908..6927
                     /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 ccgggctcgt taagacacca ggaaactcga gtttcctggt gtcttaacga
     2521 gctttttgaa ttctcgacct cgagacaaat ggcagtattc atccacaatt ttaaaagaaa
     2581 aggggggatt ggggggtaca gtgcagggga aagaatagta gacataatag caacagacat
     2641 acaaactaaa gaattacaaa aacaaattac aaaaattcaa aattttcggg tttattacag
     2701 ggacagcaga gatccacttt ggccgcggct cgagggggtt ggggttgcgc cttttccaag
     2761 gcagccctgg gtttgcgcag ggacgcggct gctctgggcg tggttccggg aaacgcagcg
     2821 gcgccgaccc tgggtctcgc acattcttca cgtccgttcg cagcgtcacc cggatcttcg
     2881 ccgctaccct tgtgggcccc ccggcgacgc ttcctgctcc gcccctaagt cgggaaggtt
     2941 ccttgcggtt cgcggcgtgc cggacgtgac aaacggaagc cgcacgtctc actagtaccc
     3001 tcgcagacgg acagcgccag ggagcaatgg cagcgcgccg accgcgatgg gctgtggcca
     3061 atagcggctg ctcagcaggg cgcgccgaga gcagcggccg ggaaggggcg gtgcgggagg
     3121 cggggtgtgg ggcggtagtg tgggccctgt tcctgcccgc gcggtgttcc gcattctgca
     3181 agcctccgga gcgcacgtcg gcagtcggct ccctcgttga ccgaatcacc gacctctctc
     3241 cccaggggga tccaccggag cttaccatga ccgagtacaa gcccacggtg cgcctcgcca
     3301 cccgcgacga cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg
     3361 ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac
     3421 tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg
     3481 cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg
     3541 gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag atggaaggcc
     3601 tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc
     3661 ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg
     3721 agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc cccttctacg
     3781 agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt
     3841 gcatgacccg caagcccggt gcctgacgcc cgccccacga cccgcagcgc ccgaccgaaa
     3901 ggagcgcacg accccatgca tcggtacctt taagaccaat gacttacaag gcagctgtag
     3961 atcttagcca ctttttaaaa gaaaaggggg gactggaagg gctaattcac tcccaacgaa
     4021 gacaagatct gctttttgct tgtactgggt ctctctggtt agaccagatc tgagcctggg
     4081 agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc
     4141 ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct
     4201 tttagtcagt gtggaaaatc tctagcagta gtagttcatg tcatcttatt attcagtatt
     4261 tataacttgc aaagaaatga atatcagaga gtgagaggaa cttgtttatt gcagcttata
     4321 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     4381 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg ctctagctat
     4441 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     4501 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     4561 attccagaag tagtgaggag gcttttttgg aggcctaggg acgtacccaa ttcgccctat
     4621 agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     4681 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     4741 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     4801 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     4861 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     4921 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     4981 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     5041 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     5101 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     5161 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     5221 aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg
     5281 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     5341 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     5401 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     5461 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     5521 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     5581 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     5641 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     5701 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     5761 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     5821 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     5881 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     5941 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     6001 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     6061 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     6121 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     6181 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     6241 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     6301 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     6361 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     6421 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     6481 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     6541 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     6601 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     6661 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     6721 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     6781 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     6841 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     6901 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     6961 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     7021 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     7081 tatc
//
