LOCUS       sequence-323516-        6797 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Acute Depletion of RBP1-EYFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6797)
  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 6797)
  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_N-RPB1_BSD_FKBPF36V_EYFP
FEATURES             Location/Qualifiers
     source          1..6797
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     127..149
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1424..1819
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             1829..1885
                     /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             1886..1909
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1919..2239
                     /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             2246..2272
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(2273..2275,2276..2986)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2315..2336)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2576..2595)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2923..2944
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(4300..4318)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4339..4355)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4339..4355)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4352..4374)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4363..4379
                     /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(4387..4393,4394..4411,4412..4417))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4432..4453
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4570..4587)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4741..5329)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4821..4840)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5500..6291,6292..6360))
                     /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     6123..6142
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6361..6465)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6533..6551
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6589..6611)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6711..6730
                     /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 ggtcgacaag tcgtctgaaa
      241 ggtgtagggt ttccttgacc ccgaattccc caagacatcc ctattctgcc ccaccgctaa
      301 tctgcttttc tcagattgtc acggtcgcgt gatctgcggc tcctgggtag cagccattct
      361 actttcagaa taggcctccg ggctcgggaa aagcttgggc tgaacatcga gcgccagggc
      421 tccgtaaagc tactagagca caggcggtgc cccaacgtcc tggggcctct ccactaataa
      481 cggctacttc caattgatag gacgcgccat cttgcctgcc ttatgcatat tcagcggtga
      541 actgaatatt catgaacgag gcccgtcccg tccctccctc cttcccccca cccccggaac
      601 ccgctccgga ggacccgaag ggccccgcct tcattaccga tgcgtaggac aaaccatttt
      661 cccgatgtgt gtggggggat actaatgaga gactttagct gaaaatgagc ctgaactccg
      721 aagctgagta aaaatggcct aactttatcc tccgttctgt aagtcctcgg tttgagtgca
      781 cgggaaaccc gaaaggagga cgacaggacc aggaacattc tcctcctcct gtcgcgtcag
      841 aaagaacacc caaccaggga gccggagccc tagcgtcaac aactccgccg cgcgcgctcc
      901 gtgtaggccg gtgcgggcgg ccccgtagcg caagggaggg cgggaaagga aggggcggga
      961 cacaagggcg aatctataaa gggcgtcact cagccagttc tctcctcaga agcgccgaga
     1021 gcgcgaccgg gacggttgga gaagaaggtg gctcccggaa gggggagaga caaactgccg
     1081 taacctctgc cgttcaggat cccggttact tatttattcg ttaccccttt tcttcttcct
     1141 ccctcccaaa acctcctcct ctcctccctt cttttgtttc ctttttggga gccgacgaat
     1201 ctccggaaag ggagaaaagg ctttctttca gcccttttcg ttcctctgcc ttcccccccc
     1261 cttcccccct cccccacctt tccctcctcc agcctttccc tccctatccc gggaggtctt
     1321 tcctggtggc cgcccggacg ggttctgagc acttaggcgg cggtggcgca ggctttttgt
     1381 agcgaggttt gcgcctgcgc agcgcgcctg cctcgccgtc gacatggcca agcctttgtc
     1441 tcaagaagaa tccaccctca ttgaaagagc aacggctaca atcaacagca tccccatctc
     1501 tgaagactac agcgtcgcca gcgcagctct ctctagcgac ggccgcatct tcactggtgt
     1561 caatgtatat cattttactg ggggaccttg tgcagaactc gtggtgctgg gcactgctgc
     1621 tgctgcggca gctggcaacc tgacttgtat cgtcgcgatc ggaaatgaga acaggggcat
     1681 cttgagcccc tgcggacggt gccgacaggt gcttctcgat ctgcatcctg ggatcaaagc
     1741 catagtgaag gacagtgatg gacagccgac ggcagttggg attcgtgaat tgctgccctc
     1801 tggttatgtg tgggagggcg gttccggagc cacgaacttc tctctgttaa agcaagcagg
     1861 agacgtggaa gaaaaccccg gtcccgatta caaggatgac gatgacaagg gttccggcgg
     1921 cgtgcaggtg gagactatct ccccaggaga cgggcgcacc ttccccaagc gcggccagac
     1981 ctgcgtggtg cactacaccg ggatgcttga agatggaaag aaagttgatt cctcccggga
     2041 cagaaacaag ccctttaagt ttatgctagg caagcaggag gtgatccgag gctgggaaga
     2101 aggggttgcc cagatgagtg tgggtcagag agccaaactg actatatctc cagattatgc
     2161 ctatggtgcc actgggcacc caggcatcat cccaccacat gccactctcg tcttcgatgt
     2221 ggagcttcta aaactggaag tcgattatcc gtacgacgta ccagactacg cagtgagcaa
     2281 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
     2341 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
     2401 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
     2461 cttcggctac ggcctgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt
     2521 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
     2581 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
     2641 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
     2701 caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt
     2761 gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca
     2821 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcta
     2881 ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     2941 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagggtg gaggtggatc
     3001 tggtggcggc ggttcaggta cccacggggg tggccccccc tccggggaca gcgcatgccc
     3061 gctgcgcacc atcaagagag tgcagttcgg agtcctgagt ccggatgaat tggtaagata
     3121 gtccgcccgt ctctctgcta tgcctttggc tgatgatcct ctgggaagat gggtcagtga
     3181 tgctgatgtt gtgcgggggt ccagccttca aaagttaaat tatgccagga cttaaacaag
     3241 cgaaatgcta tgcctgagat gagtgcttga ctggacagag aagaaagatg tttccatact
     3301 tgagcattgc tgcggttcaa cgagacaggg ttgaaagttg tacaggaagt ggatccccag
     3361 ttttagtcct ttcaccaata ttgagcatcc tgctatgtga cagtaatcag acactaggaa
     3421 cacagctgca caataaatat aacagaaact ctaccttcta aaacactatg ggaaaaacag
     3481 agaagagagc ttaagaacaa aaagaaatga gcaaaaataa gtacctagat taaagcaaaa
     3541 tgaggaaatg ctgtaagaag tcactgaaat tgaatggtgt cagggtttgg aagacatact
     3601 tgccagggat ggtgacattc ccaagtaata tcagtcatca gagctatttt tttaggtgga
     3661 ggaaacaagt gtcatgggac tgagagggaa ataaacttgt attctagaga ctgaaaagcc
     3721 agtgttgggg caggcagggt tagtgggtgc ggagagtgca ctgaagaagc acgggatatt
     3781 gtaaattacg ctctagagag tccaaatttc tgtttaaagt aggaaaccac tgggtaatta
     3841 taaaggtgac agtagtaact tatacacaca cgcatgtaca gaagctccgt aaagagctga
     3901 tgtgaagtgg gtgaggatag acaagggata gcttttggag acttggactt cttcctcctg
     3961 agaggaatag agagtggaat taaacagctg attctagact gaggcagtcc tgttggggta
     4021 agggctcaac tatagcagaa ctcaaaagaa aaaacagaaa aacctacatc agggctggta
     4081 agatggctca gcaggtaaga gaactgactg ctcttccgaa agtcatgagt tcaaatccca
     4141 gcaaccatat ggtggctcac aaccatctgt aatgagatct gatgcccttt tctggagtgt
     4201 ctgaagtcag ctacagtggg taccctcgag gtcgatcgac ggtatcgata agcttgatct
     4261 agagcggccg ccaccgcggt ggagctccag cttttgttcc ctttagtgag ggttaatttc
     4321 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     4381 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     4441 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     4501 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     4561 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     4621 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     4681 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     4741 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     4801 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     4861 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     4921 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     4981 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     5041 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     5101 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     5161 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     5221 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     5281 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     5341 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     5401 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     5461 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     5521 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     5581 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     5641 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     5701 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     5761 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     5821 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     5881 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     5941 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     6001 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     6061 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     6121 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     6181 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     6241 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     6301 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     6361 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     6421 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     6481 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     6541 tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     6601 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     6661 tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga
     6721 gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag
     6781 aaaataccgc atcaggc
//
