LOCUS       sequence-235271-        8571 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of PNRC1-W300A mutant C-MYC-FLAG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8571)
  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 8571)
  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: pLenti C-MYC-DDK Puro PNRC1-W300A
FEATURES             Location/Qualifiers
     source          1..8571
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        237..616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        618..816
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             834..1014
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1061..1186
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1683..1916
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2101..2145
                     /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"
     misc_feature    2443..2560
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        2643..3022
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3023..3226
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3252..3271
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3252..3270
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4357..4386
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4405..4428
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    4462..5035
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    4485..5037
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4629..4646)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4856..4875
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             5037..5636
                     /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(5037..5056)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5533..5553
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    complement(5666..5699)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    5755..6343
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5808..5828)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6226..6237)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(6346..6362)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6347..6363)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             6872..7052
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(7114..7702)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7194..7213)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7779..8438)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     8353..8372
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(8439..8543)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat cgcgttgaca
      241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      781 cggtaggcgt gtacggtggg aggtctatat aagcagcgcg ttttgcctgt actgggtctc
      841 tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta
      901 agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact
      961 ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtggcg
     1021 cccgaacagg gacttgaaag cgaaagggaa accagaggag ctctctcgac gcaggactcg
     1081 gcttgctgaa gcgcgcacgg caagaggcga ggggcggcga ctggtgagta cgccaaaaat
     1141 tttgactagc ggaggctaga aggagagaga tgggtgcgag agcgtcagta ttaagcgggg
     1201 gagaattaga tcgcgatggg aaaaaattcg gttaaggcca gggggaaaga aaaaatataa
     1261 attaaaacat atagtatggg caagcaggga gctagaacga ttcgcagtta atcctggcct
     1321 gttagaaaca tcagaaggct gtagacaaat actgggacag ctacaaccat cccttcagac
     1381 aggatcagaa gaacttagat cattatataa tacagtagca accctctatt gtgtgcatca
     1441 aaggatagag ataaaagaca ccaaggaagc tttagacaag atagaggaag agcaaaacaa
     1501 aagtaagacc accgcacagc aagcggccgg ccgctgatct tcagacctgg aggaggagat
     1561 atgagggaca attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta
     1621 ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga
     1681 ataggagctt tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca
     1741 atgacgctga cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat
     1801 ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag
     1861 cagctccagg caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg
     1921 atttggggtt gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg
     1981 agtaataaat ctctggaaca gatttggaat cacacgacct ggatggagtg ggacagagaa
     2041 attaacaatt acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa
     2101 aagaatgaac aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac
     2161 ataacaaatt ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt
     2221 ttaagaatag tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca
     2281 ttatcgtttc agacccacct cccaaccccg aggggacccg acaggcccga aggaatagaa
     2341 gaagaaggtg gagagagaga cagagacaga tccattcgat tagtgaacgg atcggcactg
     2401 cgtgcgccaa ttctgcagac aaatggcagt attcatccac aattttaaaa gaaaaggggg
     2461 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac
     2521 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag
     2581 cagagatcca gtttggttag taccgggccc gctctagaca tgtccaatat gaccgccatg
     2641 ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
     2701 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     2761 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     2821 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
     2881 tcaagtgtat catatgccaa gtccgccccc tattgacgtc aatgacggta aatggcccgc
     2941 ctggcattat gcccagtaca tgaccttacg ggactttcct acttggcagt acatctacgt
     3001 attagtcatc gctattacca tggtgatgcg gttttggcag tacaccaatg ggcgtggata
     3061 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
     3121 ttggcaccaa aatcaacggg actttccaaa atgtcgtaat aaccccgccc cgttgacgca
     3181 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctcgtt tagtgaaccg
     3241 tcagaatttt gtaatacgac tcactatagg gcggccggga attcgtcgac tggatccggt
     3301 accgaggaga tctgccgccg cgatcgccgg cgcgccagat ctcaagctta actagctagc
     3361 ggaccgacgc gtatgactgt cgtctccgtc ccgcagcggg agccgctcgt cctgggtggc
     3421 cgccttgcgc cgcttggctt ttcctcccga ggttactttg gggccctccc gatggtgacc
     3481 acggctccgc ctcctttacc ccggatcccg gacccccggg cactgccccc gaccctcttc
     3541 ctccctcatt tcctaggggg agatggcccg tgtctgaccc cccagcctcg cgctccagca
     3601 gctctgccca accgcagcct cgccgtggcg ggaggcactc ctcgggcagc gccgaagaag
     3661 cggcgaaaga agaaggtgcg ggccagcccc gcagggcagc tgcccagccg cttccaccag
     3721 taccagcagc accggccgag tctggagggc ggccggagcc ccgcgaccgg cccgagcgga
     3781 gcgcaggagg tcccgggccc ggccgccgcc ttggccccga gtcctgcagc cgcagccggc
     3841 acggagggag ccagccccga ccttgccccg ctgcggcccg cggctcccgg ccaaaccccc
     3901 ctcaggaaag aggttttaaa atcaaagatg ggaaaatcgg agaaaattgc ccttccccat
     3961 ggccagcttg ttcatggtat acacttgtat gagcaaccaa agataaacag acagaaaagc
     4021 aaatataact tgccactaac caagatcacc tctgcaaaaa gaaatgaaaa caacttttgg
     4081 caggattctg tttcatctga cagaattcag aagcaggaaa aaaagccttt taaaaatacc
     4141 gagaacatta aaaattcgca tttgaagaaa tcagcatttc taactgaagt gagccaaaag
     4201 gaaaattatg ctggggcaaa gtttagtgat ccaccttctc ctagtgttct tccaaagcct
     4261 cctagtcacg cgatgggaag cactgttgaa aattccaacc aaaacaggga gctgatggca
     4321 gtacacttaa aaacgctcct caaagttcaa actctcgagc agaaactcat ctcagaagag
     4381 gatctggcag caaatgatat cctggattac aaggatgacg acgataaggt ttaaacctag
     4441 gcgtagcggc cgcaaattcc gcccctctcc ctcccccccc cctaacgtta ctggccgaag
     4501 ccgcttggaa taaggccggt gtgcgtttgt ctatatgtta ttttccacca tattgccgtc
     4561 ttttggcaat gtgagggccc ggaaacctgg ccctgtcttc ttgacgagca ttcctagggg
     4621 tctttcccct ctcgccaaag gaatgcaagg tctgttgaat gtcgtgaagg aagcagttcc
     4681 tctggaagct tcttgaagac aaacaacgtc tgtagcgacc ctttgcaggc agcggaaccc
     4741 cccacctggc gacaggtgcc tctgcggcca aaagccacgt gtataagata cacctgcaaa
     4801 ggcggcacaa ccccagtgcc acgttgtgag ttggatagtt gtggaaagag tcaaatggct
     4861 ctcctcaagc gtattcaaca aggggctgaa ggatgcccag aaggtacccc attgtatggg
     4921 atctgatctg gggcctcggt gcacatgctt tacatgtgtt tagtcgaggt taaaaaaacg
     4981 tctaggcccc ccgaaccacg gggacgtggt tttcctttga aaaacacgat gataatatga
     5041 ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtcccccgg gcagtacgca
     5101 ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat ccagaccgcc
     5161 acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg
     5221 gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg ccggagagcg
     5281 tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg agcggttccc
     5341 ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc aaggagcccg
     5401 cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg
     5461 ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga
     5521 cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg
     5581 tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt gcctgatgta
     5641 caagtaggat tcgtcgaggg acctaataac ttcgtatagc atacattata cgaagttata
     5701 catgtttaag ggttccggtt ccactaggta caattcgata tcaagcttat cgataatcaa
     5761 cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt
     5821 acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct
     5881 ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc
     5941 gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg
     6001 ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc
     6061 acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc
     6121 actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctgt
     6181 gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca
     6241 gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt
     6301 cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcatcgata ccgtcgacct
     6361 cgatcgagac ctagaaaaac atggagcaat cacaagtagc aatacagcag ctaccaatgc
     6421 tgattgtgcc tggctagaag cacaagagga ggaggaggtg ggttttccag tcacacctca
     6481 ggtaccttta agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga
     6541 aaagggggga ctggaagggc taattcactc ccaacgaaga caagatatcc ttgatctgtg
     6601 gatctaccac acacaaggct acttccctga ttggcagaac tacacaccag ggccagggat
     6661 cagatatcca ctgacctttg gatggtgcta caagctagta ccagttgagc aagagaaggt
     6721 agaagaagcc aatgaaggag agaacacccg cttgttacac cctgtgagcc tgcatgggat
     6781 ggatgacccg gagagagaag tattagagtg gaggtttgac agccgcctag catttcatca
     6841 catggcccga gagctgcatc cggactgtac tgggtctctc tggttagacc agatctgagc
     6901 ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg
     6961 agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag
     7021 acccttttag tcagtgtgga aaatctctag cagcatgtga gcaaaaggcc agcaaaaggc
     7081 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     7141 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     7201 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     7261 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     7321 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     7381 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     7441 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     7501 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     7561 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     7621 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     7681 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     7741 gtggaacgaa aactcacgtt aagggatttt ggtcatgatt acgccccgcc ctgccactca
     7801 tcgcagtact gttgtaattc attaagcatt ctgccgacat ggaagccatc acaaacggca
     7861 tgatgaacct gaatcgccag cggcatcagc accttgtcgc cttgcgtata atatttgccc
     7921 atggtgaaaa cgggggcgaa gaagttgtcc atattggcca cgtttaaatc aaaactggtg
     7981 aaactcaccc agggattggc tgagacgaaa aacatattct caataaaccc tttagggaaa
     8041 taggccaggt tttcaccgta acacgccaca tcttgcgaat atatgtgtag aaactgccgg
     8101 aaatcgtcgt ggtattcact ccagagcgat gaaaacgttt cagtttgctc atggaaaacg
     8161 gtgtaacaag ggtgaacact atcccatatc accagctcac cgtctttcat tgccatacgg
     8221 aactccggat gagcattcat caggcgggca agaatgtgaa taaaggccgg ataaaacttg
     8281 tgcttatttt tctttacggt ctttaaaaag gccgtaatat ccagctgaac ggtctggtta
     8341 taggtacatt gagcaactga ctgaaatgcc tcaaaatgtt ctttacgatg ccattgggat
     8401 atatcaacgg tggtatatcc agtgattttt ttctccatac tcttcctttt tcaatattat
     8461 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     8521 aataaacaaa taggggtccc gcgcacattt ccccgaaaag tgccacctga c
//
