LOCUS       sequence-323477-        6700 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Acute Depletion of SPT5-EYFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6700)
  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 6700)
  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-SPT5_EYFP_FKBPF36V_Neo
FEATURES             Location/Qualifiers
     source          1..6700
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     127..149
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             911..934
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             944..1264
                     /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             1271..1297
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(1298..1300,1301..2011)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1340..1361)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1601..1620)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1948..1969
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2021..2077
                     /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             2072..2872
                     /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(2132..2151)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2742..2761
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     promoter        complement(4203..4221)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4242..4258)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4242..4258)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4255..4277)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4266..4282
                     /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(4290..4296,4297..4314,4315..4320))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4335..4356
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4473..4490)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4644..5232)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4724..4743)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5403..6194,6195..6263))
                     /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     6026..6045
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6264..6368)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6436..6454
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6492..6514)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6614..6633
                     /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 ggtcgacacc actgatatcc
      241 aggtgaaggt tcgagacacc tatctggaca cacaaatagt ggggcagaca ggcgtcatcc
      301 gcagtgtcac ggtaagtgtg gccctaggag gtagacatgt gtacccctct accccccagc
      361 ctttgtagtt atttttatct ctgttgctgt ggatggaacc ctaggcctca cacatgctag
      421 gctagtgcct accactgaac tacatcccca acttattgtt atagttttag atttaacttt
      481 acaactataa tgcatggtaa gctgtgatgg acaagcagat gtggaaggct ggggtgtctg
      541 aggcagctgg agggtgggca gtaagagcag gctgtccgtc cttctgcagg gaggcatgtg
      601 ctccgtgtac ctgaaggaca gtgagaaggt ggtcagcatc tccagtgaac atctggagcc
      661 catcacgccc accaagaaca acaaggtaga gatggggcct tgggaggata ccggagggaa
      721 tgacagggct gctgctgggc ctgacactgg ctgtcgcctc caggtgaagg tgattctggg
      781 tgaggaccga gaggctaccg gggtcctact gagcattgat ggtgaggatg gcatcatccg
      841 catggaccta gaggaccagc agatcaagat tctaaacctc cgcttcctag ggaagctcct
      901 ggaagtcgac gattacaagg atgacgatga caagggttcc ggcggcgtgc aggtggagac
      961 tatctcccca ggagacgggc gcaccttccc caagcgcggc cagacctgcg tggtgcacta
     1021 caccgggatg cttgaagatg gaaagaaagt tgattcctcc cgggacagaa acaagccctt
     1081 taagtttatg ctaggcaagc aggaggtgat ccgaggctgg gaagaagggg ttgcccagat
     1141 gagtgtgggt cagagagcca aactgactat atctccagat tatgcctatg gtgccactgg
     1201 gcacccaggc atcatcccac cacatgccac tctcgtcttc gatgtggagc ttctaaaact
     1261 ggaagtcgat tatccgtacg acgtaccaga ctacgcagtg agcaagggcg aggagctgtt
     1321 caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag
     1381 cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg
     1441 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccttcg gctacggcct
     1501 gcagtgcttc gcccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat
     1561 gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac
     1621 ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat
     1681 cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca
     1741 caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg
     1801 ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat
     1861 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agctaccagt ccgccctgag
     1921 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg
     1981 gatcactctc ggcatggacg agctgtacaa gggatccggc gcaacaaact tctctctgct
     2041 gaaacaagcc ggagatgtcg aagagaatcc tggaccgatg attgaacaag atggattgca
     2101 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
     2161 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
     2221 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc
     2281 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
     2341 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc
     2401 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
     2461 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
     2521 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
     2581 cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca
     2641 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     2701 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     2761 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     2821 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gaggtacccg
     2881 cccaagtgtg cagtgcagac atggagccag cagcccctct ggccttgttt ggcctagcat
     2941 ggaaccctga gtctggagca agccagggca atcactctgt atttcctttt cctccttccc
     3001 tgatactcta ggagtcagag cagagtctga ggaatgttcc tgccttcttc tgcagcccca
     3061 gcttgacttt gtgttcaggt ctttcaataa agagaagctg tttgaaaagt gtgtgtacga
     3121 agagttttct tggtcttaag tatagatatg aagttgggga aataaccaga tcttaagtaa
     3181 gtttctccat ctagaaaaat aatgcttctg gaactccaaa gggctcttca gagtctgttg
     3241 cggacaggag aaagatgatt agggctggcc tttgtcattc aggcctgtca cccaagctac
     3301 ttagacctgg ggaggacgat cacaggttca tgtcttatct gagcaagtct cactgagggc
     3361 aacaagcccc tttactgcag tgcttcttaa cctgggggga gtaaacatgt ttgggggagt
     3421 cgaacgatcc tttcacagga gtcacatcag atatcctgta tatcagatat ttacattatg
     3481 tttcataact agcaaaatta cagttatgaa gtagcaatga aaatactttt atggttgggg
     3541 atcattacaa cataaggaac tgtgttaaag ggttgcagta tatggttaag aactgcctta
     3601 ggacccgagc ctcagagggc agggcacagt ggtacatgcc tttgatccta gcatttggga
     3661 ggcagaggct ggtgggtctc tgtgctgtct acagagtgga ttctaggaca gtcaaagcta
     3721 aatagtaaga ccctatgggg ggtgggggtg gacaggtata ctcagcaatt cagctgggac
     3781 acagcttaat gaagagtgct ttacaaagta ggtgtggcac catggcttta gaaagccagt
     3841 ctgtaaaaag taacaggtaa gaatggagtt tgggggctgg agagatggct cagcggttaa
     3901 gagtactggc tgttcttcca gagggtctga gttcaatcta taaagagata tgatcccctc
     3961 ttctggcatg caattgtacc tgcagacagt gcamtcatac atttaaaatg agtgaatgaa
     4021 cgagtgaata ataaatgatt aaacaaataa aggaatgaat aaacaaacaa atagaaaaca
     4081 acttgggcaa gactcatggt ggctcccgca cccctaattc aggtaccctc gaggtcgatc
     4141 gacggtatcg ataagcttga tctagagcgg ccgccaccgc ggtggagctc cagcttttgt
     4201 tccctttagt gagggttaat ttcgagcttg gcgtaatcat ggtcatagct gtttcctgtg
     4261 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     4321 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     4381 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     4441 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     4501 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     4561 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     4621 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     4681 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     4741 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     4801 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     4861 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     4921 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     4981 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     5041 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     5101 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     5161 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     5221 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     5281 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     5341 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     5401 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     5461 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     5521 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     5581 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     5641 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     5701 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     5761 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     5821 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     5881 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     5941 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     6001 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     6061 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     6121 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     6181 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     6241 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     6301 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     6361 gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg
     6421 acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat
     6481 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     6541 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     6601 tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa
     6661 ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc
//
