LOCUS       sequence-489758-        6197 bp    DNA     circular SYN 07-NOV-2025
DEFINITION  PHF1 KO donor plasmid with puromycin resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6197)
  AUTHORS   Much C, Rajkumar SM, Chen L, Cohen JM, Gade AR, Pitt GS, Long Y
  TITLE     Distinct specificity and functions of PRC2 subcomplexes in human 
            stem cells and cardiac differentiation.
  JOURNAL   Mol Cell. 2025 Aug 7:S1097-2765(25)00615-X. doi: 
            10.1016/j.molcel.2025.07.014.
  PUBMED    40803325
REFERENCE   2  (bases 1 to 6197)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 7, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Cell. 
            2025 Aug 7:S1097-2765(25)00615-X. doi: 
            10.1016/j.molcel.2025.07.014."
COMMENT     Sequence Label: pUC19_PHF1_KO_Puro_donor
FEATURES             Location/Qualifiers
     source          1..6197
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    1172..1293
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    1294..1415
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    1416..1537
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    1540..1651
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    1754..1787
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1788..2298
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     2217..2235
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             2308..2907
                     /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(2308..2327)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2804..2824
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    2943..3167
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    3168..3201
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(3976..3992)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3976..3992)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3989..4011)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4000..4016
                     /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)."
     promoter        complement(join(4024..4030,4031..4048,4049..4054))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4069..4090
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4207..4224)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4378..4966)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4458..4477)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5137..5928,5929..5997))
                     /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     5760..5779
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5998..6102)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6170..6188
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctctgg aagctgggaa
      421 actagcaact aggtttgggg tcggggggag ggctgttagt taccgagagg agagccagtc
      481 actagagttg gaagaggact caaaaagctg gtgatgggtg cccagtggtt tcctatcaag
      541 gtgtctcagg gatttggaga tgggggcaca gatacttcat tcttgatggc aaaataggca
      601 aagaaaatgg ggaggctggc cgggcccggt ggctcacgcc tgtaatccca gcactttggg
      661 aagctgaggt gggtggatca tgaggtcagg agttcgagac cagtttggcc aacatggcca
      721 acacctcatc tctactaaaa atacaaaaat tagccaggca tggtggcatg cgcctgtaat
      781 cccacctact tgggagactg aggcaggaga attgcctgaa cccgggaggt ggaggttgca
      841 gtgagccaag attgtgccac tgcactctgg cctgggcgac agagcaaaac tccatctcag
      901 gaaaaaaaaa aaaaaaagaa aaagaaaatg gggaaggttt ctcagcattc ataaccttct
      961 cctccccatt tcttttctgg ctaggccccc ccaggatgca atggcgcagc ccccccggct
     1021 gagccgctct ggtgcctcct cactttggga cccagcttct cctgctccca cctctggccc
     1081 caggcctcgg ctttgggagg gtcaagatgt gctggccaga tggactgatg ggctgctata
     1141 cttgggtacc acctttgaac aatcaggtac caacttgttt attgcagctt ataatggtta
     1201 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     1261 ttgtggtttg tccaaactca tcaatgtatc ttaaacttgt ttattgcagc ttataatggt
     1321 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     1381 agttgtggtt tgtccaaact catcaatgta tcttaaactt gtttattgca gcttataatg
     1441 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     1501 ctagttgtgg tttgtccaaa ctcatcaatg tatcttagac tgtgccttct agttgccagc
     1561 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     1621 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     1681 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagagaat agcaggcatg
     1741 cggccgcgat atcataactt cgtataatgt atgctatacg aagttatggg gttggggttg
     1801 cgccttttcc aaggcagccc tgggtttgcg cagggacgcg gctgctctgg gcgtggttcc
     1861 gggaaacgca gcggcgccga ccctgggtct cgcacattct tcacgtccgt tcgcagcgtc
     1921 acccggatct tcgccgctac ccttgtgggc cccccggcga cgcttcctgc tccgccccta
     1981 agtcgggaag gttccttgcg gttcgcggcg tgccggacgt gacaaacgga agccgcacgt
     2041 ctcactagta ccctcgcaga cggacagcgc cagggagcaa tggcagcgcg ccgaccgcga
     2101 tgggctgtgg ccaatagcgg ctgctcagca gggcgcgccg agagcagcgg ccgggaaggg
     2161 gcggtgcggg aggcggggtg tggggcggta gtgtgggccc tgttcctgcc cgcgcggtgt
     2221 tccgcattct gcaagcctcc ggagcgcacg tcggcagtcg gctccctcgt tgaccgaatc
     2281 accgacctct ctccccagca attcaccatg accgagtaca agcccacggt gcgcctcgcc
     2341 acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc
     2401 gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa
     2461 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc
     2521 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc
     2581 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc
     2641 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg
     2701 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc
     2761 gagcgcgccg gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac
     2821 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg
     2881 tgcatgaccc gcaagcccgg tgcctgactc gagtctagac caattggttt aaaccctgca
     2941 ggctgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     3001 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     3061 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     3121 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggata acttcgtata
     3181 atgtatgcta tacgaagtta tcctagggta agaccttcta cctctgacct tcttcctagt
     3241 tcccttatct aattctggtt cccatttcat cctgtttgct cacccattcc aggtggacag
     3301 tgctagggag gtgtgtctgg tccagtttga ggatgattcg cagtttctgg ttctatggaa
     3361 agacattagc cctggtaaga ctctagagac ctgagattgc acatcccatg gaaaacaaac
     3421 ctggcctaga ggggcagaaa gagacttaaa ggcaggccct gtgacactgt gttctctcac
     3481 agctgccctc cctggagagg aactcctctg ttgtgtctgt cgctctgaga ctgtggtccc
     3541 tgggaaccgg ctggtcagct gtgagaagtg tcgccatggt gagagggcag gtcacctgaa
     3601 tggtccagct tgctcttccc tccaggatgg tctctatatc acctgtcctg gccttagtcc
     3661 ccaagccact gctctggcca ggctgcttag ccttatctta gtcctcatcc gctttcagcc
     3721 cagggagaag cagccatgga aaggggtggt ataacctttg caggcagaag atggtttaaa
     3781 tgcatccttt tctaaccata tgacctcgga caagtccctt aacctctagg agcctcagtt
     3841 ccctgacttg taaaataagg ataatgcacc ccctcatcaa gaccatggtc agggattaaa
     3901 tgagatgggt aaagactatt cctgtcccaa taccaagcac acacaggtat gcaataaaag
     3961 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     4021 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     4081 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     4141 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     4201 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     4261 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     4321 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     4381 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     4441 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     4501 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     4561 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     4621 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     4681 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     4741 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     4801 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4861 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4921 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4981 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     5041 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     5101 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     5161 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     5221 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     5281 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     5341 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     5401 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     5461 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     5521 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     5581 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     5641 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     5701 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     5761 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     5821 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     5881 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     5941 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     6001 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     6061 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6121 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     6181 cacgaggccc tttcgtc
//
