LOCUS       sequence-323501-        7546 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Acute Depletion of WAPL-EYFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7546)
  AUTHORS   Zhu Y, Denholtz M, Lu H, Murre C
  TITLE     Calcium signaling instructs NIPBL recruitment at active enhancers 
            and promoters via distinct mechanisms to reconstruct genome 
            compartmentalization.
  JOURNAL   Genes Dev. 2021 Jan 1;35(1-2):65-81. doi: 10.1101/gad.343475.120. 
            Epub 2020 Dec 17.
  PUBMED    33334824
REFERENCE   2  (bases 1 to 7546)
  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: "Genes 
            Dev."; date: "2021-01-1"; volume: "35(1-2):65-81. doi"; pages: " 
            10.1101/gad.343475.120. Epub 2020 Dec 17"
COMMENT     Sequence Label: pFlox c-WAPL_EYFP_FKBPF36V_Neo
FEATURES             Location/Qualifiers
     source          1..7546
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     127..149
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1259..1282
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1292..1612
                     /codon_start=1
                     /product="F36V mutant of FK506-binding protein FKBP12"
                     /label=DmrB
                     /note="binds synthetic ligands that do not bind wild-type 
                     FKBP"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             1619..1645
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(1646..1648,1649..2359)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1688..1709)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1949..1968)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2296..2317
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2369..2425
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2420..3220
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="GPMIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQG
                     RPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
                     LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEE
                     HQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQD
                     IALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2480..2499)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3090..3109
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     promoter        complement(5049..5067)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5088..5104)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5088..5104)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5101..5123)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5112..5128
                     /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(5136..5142,5143..5160,5161..5166))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5181..5202
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5319..5336)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5490..6078)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5570..5589)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6249..7040,7041..7109))
                     /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     6872..6891
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7110..7214)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7282..7300
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7338..7360)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7460..7479
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc
       61 tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag
      121 ggttttccca gtcacgacgt tgtaaaacga cggccgcggc ggccgcacta gtggccgcgg
      181 cggccgcact agtggccgcg gcggccgcag cttgcctgca ggtcgactag gtaagtgaat
      241 gtggactgga aatgacgtat gaattttggg gtcctttgta gatgactacg ggatatgtag
      301 cagagccaac agcatcaagt gagttacagt atcatttttt accagcttga ctgtttttaa
      361 tctgtttctt cagatcaatg taactacagt aagggaatat cttccagaag gagatttctc
      421 cataatgaca gagatgctta aaaagttctt aagcttcatg aatcttacgg taagtaactt
      481 ttattgtata ttgcagttta tatgagtttt tgattatttt ccagacaaag tcttaacact
      541 gtagcccagg ctggcaatcc acatgtttca gccattggac cagtgggacc tggctagttt
      601 atgtgttttt caatttaaat gtaatacatc gaaggtgaaa ataataatcc catagtgttg
      661 ttcttccctg tggcaaaata gttaacataa gcagcttaca gaaggtacat tttggtttat
      721 agttctaaat ttttgtgtgt gtgttttttt ttctgtttgt ttttgttttt ttgagacagg
      781 gtttctctgt gtagccctgg ctgtcctgga actcaccctg tagaccaggc tggcctcgaa
      841 ctcagaaatc cgcctgcttc tgcctctcaa gtgctaggat caaaggcgtg cgccaccacc
      901 accaggctag ttctaaaatt tttagtcatg tgtggcaaga gcagcgtgga agagcagagt
      961 ggccagctaa tagagaatgc ctgtgtatct gctttctcct tttattccag ctgtgcttct
     1021 aggctgtggg atgatgccac ccacaaacag aaaactattg tggcagaatg ttatattact
     1081 aaggcaatga tttaatttca gtgtactctt tactacgtag actttgatac ttttttttgt
     1141 ttaccatttt gttaagtatt aactctaatt tctctccttc ccctcttagt gtgctgttgg
     1201 aacaacaggc cagaagtcta tctctagagt gattgaatat ttggaacatt gcgtcgacga
     1261 ttacaaggat gacgatgaca agggttccgg cggcgtgcag gtggagacta tctccccagg
     1321 agacgggcgc accttcccca agcgcggcca gacctgcgtg gtgcactaca ccgggatgct
     1381 tgaagatgga aagaaagttg attcctcccg ggacagaaac aagcccttta agtttatgct
     1441 aggcaagcag gaggtgatcc gaggctggga agaaggggtt gcccagatga gtgtgggtca
     1501 gagagccaaa ctgactatat ctccagatta tgcctatggt gccactgggc acccaggcat
     1561 catcccacca catgccactc tcgtcttcga tgtggagctt ctaaaactgg aagtcgatta
     1621 tccgtacgac gtaccagact acgcagtgag caagggcgag gagctgttca ccggggtggt
     1681 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga
     1741 gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa
     1801 gctgcccgtg ccctggccca ccctcgtgac caccttcggc tacggcctgc agtgcttcgc
     1861 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
     1921 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
     1981 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     2041 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat
     2101 catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga
     2161 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
     2221 cgtgctgctg cccgacaacc actacctgag ctaccagtcc gccctgagca aagaccccaa
     2281 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg
     2341 catggacgag ctgtacaagg gatccggcgc aacaaacttc tctctgctga aacaagccgg
     2401 agatgtcgaa gagaatcctg gaccgatgat tgaacaagat ggattgcacg caggttctcc
     2461 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     2521 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     2581 cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac
     2641 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     2701 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     2761 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     2821 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     2881 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     2941 caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     3001 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     3061 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     3121 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     3181 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga ggtaccgctt caggtgcttg
     3241 gtaatgctga agctatcctt agacaaagaa aagacatgaa agaagtcctt gaagatatac
     3301 caagaacatt catcagtatc attcattcgt gtttggattt ttatgatcac ccgatttctt
     3361 catcatgcat tctgcgtttg ctaaatgaca gttactacat caatctgcag ctatcaaaaa
     3421 tgagggaaaa ggttcaggct gttaacaatc ccatgcagta tttaaataca cttacttggc
     3481 agagtctata cctgcccctg tttccttgct gtatttcagg caaggtttaa agagggcaat
     3541 ttgtgctgct gttagtgcaa ctgctgtgta tgttgagcca ctgttgtcat gccagtcagg
     3601 tgcaaaggca gcttagctac tgaggtattg aatgttctga ggacattcta gacaacagct
     3661 tagatccttt ttcaggctca tttgcttttg ctttttgttg aatgattcca atcgtaaata
     3721 aagcttttaa taattttgtg aattttttgg ttgttgttcc ctgaactact gtctatattt
     3781 aaaatgagat ggaatccaaa gacacaaggg attaatagta tattttttta ttcttgatta
     3841 ggtttgggtg ttgaactatt tttcacttct gagaccatga ccatattcaa tgtcatacca
     3901 ttatgtgtca tagctatagg cacaagaaaa caaatagcag tttgagggaa tattatgtta
     3961 tataagataa tgtgccctgt taaaggatta agcaaaaata gacaaaccca gggtagttta
     4021 cacttaatgc tggggaggct cttgaaacat tgttacgttt tgagggacgt tctctagata
     4081 tattttctaa tgttcagttc agtaaatata ataaggaagc taaaacacca atgtggaatc
     4141 cgtttttcca gatagcgtgt atattcttct atagagtgac aggatcaatt gcataagcgc
     4201 aaagccttaa aattgctggt gtagagaaga ccctttttta ttcagattct atgttcatag
     4261 agcaattgtt tagaaaatgg ttgtggaaca cacagaacat ggtttatagg gtaaatttca
     4321 tagattgttt tttctttttt tttttttctt tttttttttt gtattgttca gaccactggt
     4381 tatacatttt ctcctttttt tattaattgg cctgaggaca ggctttccag agatgtttca
     4441 ttaatacttt taattacctt tcagatagtt acatcatgtt tcttcattgg atttgtaaaa
     4501 cttaaagcca taaaaatatt agtttggtgt gtattgggga aaatagctaa aagtctaact
     4561 ttgactcatt ttagactttg ttatttcctt gtataaagta acaaaccggg gctcttgtat
     4621 ccgtgccagc tgtaatgttt ttaaatgcag tggctgcctt ctattgtctt cctatttttg
     4681 ataatgcaga ttgttgggaa atctataagg aagtaactga ctctaggcaa attgttttct
     4741 tcctcctacc cgcccaaccc cctacccgtc cgtcaccttt taggaacaca gtatgccagt
     4801 gtaatgtggg aaccattgag actgtatttg ccctgagtac caaactaatt tagtgaattt
     4861 ttttttaaac atactggtaa atgatgtcca caaatttcct tagatatatt ttaattaatt
     4921 gtcttaaaat acattgatat ttattaatca ttggatttgg ggacagaggt accctcgagg
     4981 tcgatcgacg gtatcgataa gcttgatcta gagcggccgc caccgcggtg gagctccagc
     5041 ttttgttccc tttagtgagg gttaatttcg agcttggcgt aatcatggtc atagctgttt
     5101 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     5161 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     5221 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     5281 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     5341 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     5401 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     5461 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     5521 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     5581 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     5641 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     5701 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     5761 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5821 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     5881 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     5941 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     6001 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     6061 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     6121 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     6181 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     6241 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     6301 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     6361 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     6421 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     6481 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     6541 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     6601 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     6661 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     6721 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     6781 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     6841 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     6901 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     6961 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     7021 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     7081 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     7141 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     7201 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt
     7261 atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc
     7321 ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg
     7381 taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt
     7441 cggggctggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg
     7501 tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggc
//
