LOCUS       sequence-494266-        6216 bp    DNA     circular SYN 02-DEC-2025
DEFINITION  PHF19 KO donor plasmid with puromycin resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6216)
  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 6216)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 2, 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_PHF19_KO_Puro_donor
FEATURES             Location/Qualifiers
     source          1..6216
                     /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    1207..1328
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    1329..1450
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    1451..1572
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    1575..1686
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    1789..1822
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1823..2333
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     2252..2270
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             2343..2942
                     /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(2343..2362)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2839..2859
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    2978..3202
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    3203..3236
                     /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(3995..4011)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3995..4011)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4008..4030)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4019..4035
                     /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(4043..4049,4050..4067,4068..4073))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4088..4109
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4226..4243)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4397..4985)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4477..4496)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5156..5947,5948..6016))
                     /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     5779..5798
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6017..6121)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6189..6207
                     /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 cgagctctaa ttgagaacag
      421 aaggggggat tacaaaagac agatgaccca gtctacacta gctacaagcc acccagtttt
      481 cctttctagg tgaattttct ctggacagat ctcaggttag ggatggttgc aggggttggg
      541 ggtaaaaaca aaacccaact gcttgcctgg agaagatcct ccatccagag cacatggact
      601 gtgtttgctt aaggaataaa acctcagcct cctgagacag ggagtggttc aggcccgctg
      661 ggtaggggac tttgttctgt gtgggttgag agatccattc tctttaaaag gtagtgtggt
      721 atgggccatg gctctgcctt tggcccatca gcctcttctc cttgccacac ctgtttccct
      781 atctgtaaga tgtgaataat aataatgtct ccctcatagg atggtcacga ggaatcagtg
      841 agataataac caagtctggc ttatagcaca gggtacagaa atggtggttt ccttccttcc
      901 cctgatgggc tttctgaagc agggaagcaa gaagcaaaaa ccctgttctt tctctcccag
      961 tgtcagggga agctgatgga gaatcgagct ctggatccag ggactcggga ctcctatggt
     1021 gccaccagcc acctccccaa caagggggcc ctggcgaagg tcaagaacaa cttcaaagac
     1081 ttgatgtcca aactgacgga gggccagtat gtgctgtgcc ggtggacaga tggcctgtac
     1141 tacctcggga agatcaattg atcttcccga ggtagtacag caagaggcac tggcggggca
     1201 gcaggcaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     1261 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     1321 gtatcttaaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     1381 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     1441 atgtatctta aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     1501 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     1561 caatgtatct tagactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     1621 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     1681 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     1741 caagggggag gattgggaag agaatagcag gcatgcggcc gcgatatcat aacttcgtat
     1801 aatgtatgct atacgaagtt atggggttgg ggttgcgcct tttccaaggc agccctgggt
     1861 ttgcgcaggg acgcggctgc tctgggcgtg gttccgggaa acgcagcggc gccgaccctg
     1921 ggtctcgcac attcttcacg tccgttcgca gcgtcacccg gatcttcgcc gctacccttg
     1981 tgggcccccc ggcgacgctt cctgctccgc ccctaagtcg ggaaggttcc ttgcggttcg
     2041 cggcgtgccg gacgtgacaa acggaagccg cacgtctcac tagtaccctc gcagacggac
     2101 agcgccaggg agcaatggca gcgcgccgac cgcgatgggc tgtggccaat agcggctgct
     2161 cagcagggcg cgccgagagc agcggccggg aaggggcggt gcgggaggcg gggtgtgggg
     2221 cggtagtgtg ggccctgttc ctgcccgcgc ggtgttccgc attctgcaag cctccggagc
     2281 gcacgtcggc agtcggctcc ctcgttgacc gaatcaccga cctctctccc cagcaattca
     2341 ccatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg
     2401 tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg
     2461 accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg
     2521 acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg
     2581 agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg
     2641 gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg
     2701 agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg
     2761 gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc
     2821 tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg
     2881 ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct
     2941 gactcgagtc tagaccaatt ggtttaaacc ctgcaggctg tgccttctag ttgccagcca
     3001 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     3061 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     3121 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
     3181 ggggatgcgg tgggctctat ggataacttc gtataatgta tgctatacga agttatccta
     3241 ggtggcccct gttctcagac tcagaatctc tctgctcatg ttccagcctg gctgtcagac
     3301 cagggggcag ggggattgct ggagaattcc cttcttgtcc ctcatgccca aagctggtgg
     3361 tgggaggggc cagagcctgg tcttgcaggc accatcccta gggtcaacat catccaacta
     3421 gttctttaga aaagtgctct gagagcctac aaaacaaaaa agtcaggttt gagaggatca
     3481 cagaggagca tctgacttgt gcttctgtcc tcaacattct gaccaagttg ccatccatcg
     3541 gtccctggct gtacgactcc aggtgactag gagtacaccc tgtccaggaa ctgccatcct
     3601 tcttcccctt ctagccctgc tttctggaac atacggagac aaaaatggag gaggaagact
     3661 atgttccagg ttggcttttc cccagccctt tctcacttgc tccttcctat ctgtacccca
     3721 ggtcagcagc tctaagcaaa gctgcctcgt gactttcgaa gataattcca aatactgggt
     3781 cctatggaag gacatacagc atggtgagtt ttcctaacat ttcgggtagc ccccaacctc
     3841 cccaggtctt ctctcctctg ctgccttcat ttcttccttc tgtccttcca tctcttactc
     3901 ctggcatctg cccttgacca tgaaccccaa gcccacctgt gatgatgatc aaatctggtt
     3961 gtatattgac ttaggaaagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     4021 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     4081 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     4141 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     4201 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     4261 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4321 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4381 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4441 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4501 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4561 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4621 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4681 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4741 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4801 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     4861 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     4921 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     4981 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     5041 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     5101 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     5161 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     5221 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     5281 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     5341 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     5401 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5461 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5521 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5581 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5641 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5701 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5761 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     5821 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     5881 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     5941 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     6001 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     6061 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     6121 gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa
     6181 cctataaaaa taggcgtatc acgaggccct ttcgtc
//
