LOCUS       sequence-320629-        7084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Suppress POLD1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7084)
  AUTHORS   Sviderskiy VO, Blumenberg L, Gorodetsky E, Karakousi TR, Hirsh N, 
            Alvarez SW, Terzi EM, Kaparos E, Whiten GC, Ssebyala S, Tonzi P, Mir
            H, Neel BG, Huang TT, Adams S, Ruggles KV, Possemato R
  TITLE     Hyperactive CDK2 Activity in Basal-like Breast Cancer Imposes a 
            Genome Integrity Liability that Can Be Exploited by Targeting DNA 
            Polymerase epsilon.
  JOURNAL   Mol Cell. 2020 Oct 28. pii: S1097-2765(20)30723-1. doi: 
            10.1016/j.molcel.2020.10.016.
  PUBMED    33152268
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: "Mol Cell. 
            2020 Oct 28. pii: S1097-2765(20)30723-1. doi: 
            10.1016/j.molcel.2020.10.016."
COMMENT     Sequence Label: pLKO.1P POLD1_1
FEATURES             Location/Qualifiers
     source          1..7084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    555..788
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             973..1017
                     /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        1315..1563
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1315..1335
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1486..1505
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    1660..1777
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1826..2336
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     2255..2273
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             2358..2957
                     /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(2358..2377)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2854..2874
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3085..3318
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3390..3511
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3427..3446)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3481..3500
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3551..3686
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3706..3725)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3707..3725)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3735..3752)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3735..3751)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3744..3766)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3893..4348
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3980..3999)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4190..4211
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4374..4478
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4479..4547,4548..5339)
                     /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(4697..4716)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5510..6098
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5999..6018
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6252..6269
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6386..6407
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6422..6427,6428..6445,6446..6452)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6460..6476
                     /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     6465..6487
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6484..6500
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6484..6500
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6519..6539
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6521..6539
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6567..6793
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6794..6974
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    join(7021..7084,1..62)
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
ORIGIN
        1 tgagtacgcc aaaaattttg actagcggag gctagaagga gagagatggg tgcgagagcg
       61 tcagtattaa gcgggggaga attagatcgc gatgggaaaa aattcggtta aggccagggg
      121 gaaagaaaaa atataaatta aaacatatag tatgggcaag cagggagcta gaacgattcg
      181 cagttaatcc tggcctgtta gaaacatcag aaggctgtag acaaatactg ggacagctac
      241 aaccatccct tcagacagga tcagaagaac ttagatcatt atataataca gtagcaaccc
      301 tctattgtgt gcatcaaagg atagagataa aagacaccaa ggaagcttta gacaagatag
      361 aggaagagca aaacaaaagt aagaccaccg cacagcaagc ggccgctgat cttcagacct
      421 ggaggaggag atatgaggga caattggaga agtgaattat ataaatataa agtagtaaaa
      481 attgaaccat taggagtagc acccaccaag gcaaagagaa gagtggtgca gagagaaaaa
      541 agagcagtgg gaataggagc tttgttcctt gggttcttgg gagcagcagg aagcactatg
      601 ggcgcagcgt caatgacgct gacggtacag gccagacaat tattgtctgg tatagtgcag
      661 cagcagaaca atttgctgag ggctattgag gcgcaacagc atctgttgca actcacagtc
      721 tggggcatca agcagctcca ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa
      781 cagctcctgg ggatttgggg ttgctctgga aaactcattt gcaccactgc tgtgccttgg
      841 aatgctagtt ggagtaataa atctctggaa cagatttgga atcacacgac ctggatggag
      901 tgggacagag aaattaacaa ttacacaagc ttaatacact ccttaattga agaatcgcaa
      961 aaccagcaag aaaagaatga acaagaatta ttggaattag ataaatgggc aagtttgtgg
     1021 aattggttta acataacaaa ttggctgtgg tatataaaat tattcataat gatagtagga
     1081 ggcttggtag gtttaagaat agtttttgct gtactttcta tagtgaatag agttaggcag
     1141 ggatattcac cattatcgtt tcagacccac ctcccaaccc cgaggggacc cgacaggccc
     1201 gaaggaatag aagaagaagg tggagagaga gacagagaca gatccattcg attagtgaac
     1261 ggatctcgac ggtatcgatc acgagactag cctcgagcgg ccgccccctt caccgagggc
     1321 ctatttccca tgattccttc atatttgcat atacgataca aggctgttag agagataatt
     1381 ggaattaatt tgactgtaaa cacaaagata ttagtacaaa atacgtgacg tagaaagtaa
     1441 taatttcttg ggtagtttgc agttttaaaa ttatgtttta aaatggacta tcatatgctt
     1501 accgtaactt gaaagtattt cgatttcttg gctttatata tcttgtggaa aggacgaaac
     1561 accggcggtt acaacatcca gaacttctcg agaagttctg gatgttgtaa ccgtttttga
     1621 attctcgacc tcgagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     1681 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     1741 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     1801 agatccactt tggccgcggc tcgagggggt tggggttgcg ccttttccaa ggcagccctg
     1861 ggtttgcgca gggacgcggc tgctctgggc gtggttccgg gaaacgcagc ggcgccgacc
     1921 ctgggtctcg cacattcttc acgtccgttc gcagcgtcac ccggatcttc gccgctaccc
     1981 ttgtgggccc cccggcgacg cttcctgctc cgcccctaag tcgggaaggt tccttgcggt
     2041 tcgcggcgtg ccggacgtga caaacggaag ccgcacgtct cactagtacc ctcgcagacg
     2101 gacagcgcca gggagcaatg gcagcgcgcc gaccgcgatg ggctgtggcc aatagcggct
     2161 gctcagcagg gcgcgccgag agcagcggcc gggaaggggc ggtgcgggag gcggggtgtg
     2221 gggcggtagt gtgggccctg ttcctgcccg cgcggtgttc cgcattctgc aagcctccgg
     2281 agcgcacgtc ggcagtcggc tccctcgttg accgaatcac cgacctctct ccccaggggg
     2341 atccaccgga gcttaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg
     2401 acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc
     2461 acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca
     2521 cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg
     2581 tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca
     2641 tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc
     2701 cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc
     2761 agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg
     2821 gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg
     2881 gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc
     2941 gcaagcccgg tgcctgacgc ccgccccacg acccgcagcg cccgaccgaa aggagcgcac
     3001 gaccccatgc atcggtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc
     3061 actttttaaa agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagatc
     3121 tgctttttgc ttgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg
     3181 gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag
     3241 tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag
     3301 tgtggaaaat ctctagcagt agtagttcat gtcatcttat tattcagtat ttataacttg
     3361 caaagaaatg aatatcagag agtgagagga acttgtttat tgcagcttat aatggttaca
     3421 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     3481 gtggtttgtc caaactcatc aatgtatctt atcatgtctg gctctagcta tcccgcccct
     3541 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     3601 actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa
     3661 gtagtgagga ggcttttttg gaggcctagg gacgtaccca attcgcccta tagtgagtcg
     3721 tattacgcgc gctcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
     3781 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     3841 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg ggacgcgccc
     3901 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     3961 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     4021 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     4081 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     4141 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     4201 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     4261 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     4321 tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa
     4381 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4441 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     4501 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     4561 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     4621 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     4681 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     4741 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     4801 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     4861 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     4921 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     4981 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     5041 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     5101 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     5161 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     5221 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     5281 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     5341 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     5401 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     5461 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     5521 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     5581 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     5641 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     5701 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     5761 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     5821 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     5881 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     5941 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     6001 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     6061 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     6121 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     6181 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     6241 cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc
     6301 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     6361 aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg
     6421 ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc
     6481 acacaggaaa cagctatgac catgattacg ccaagcgcgc aattaaccct cactaaaggg
     6541 aacaaaagct ggagctgcaa gcttaatgta gtcttatgca atactcttgt agtcttgcaa
     6601 catggtaacg atgagttagc aacatgcctt acaaggagag aaaaagcacc gtgcatgccg
     6661 attggtggaa gtaaggtggt acgatcgtgc cttattagga aggcaacaga cgggtctgac
     6721 atggattgga cgaaccactg aattgccgca ttgcagagat attgtattta agtgcctagc
     6781 tcgatacata aacgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta
     6841 actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg
     6901 tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg
     6961 gaaaatctct agcagtggcg cccgaacagg gacttgaaag cgaaagggaa accagaggag
     7021 ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg caagaggcga ggggcggcga
     7081 ctgg
//
