LOCUS       sequence-375869-        7413 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cell line generation via recombination-mediated cassette exchange 
            (RMCE) and stable expression of MBD1.MBD-R22A-13XL-BASU.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7413)
  AUTHORS   Villasenor R, Pfaendler R, Ambrosi C, Butz S, Giuliani S, Bryan E, 
            Sheahan TW, Gable AL, Schmolka N, Manzo M, Wirz J, Feller C, von 
            Mering C, Aebersold R, Voigt P, Baubec T
  TITLE     ChromID identifies the protein interactome at chromatin marks.
  JOURNAL   Nat Biotechnol. 2020 Mar 2. pii: 10.1038/s41587-020-0434-2. doi: 
            10.1038/s41587-020-0434-2.
  PUBMED    32123383
REFERENCE   2  (bases 1 to 7413)
  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: "Nat 
            Biotechnol. 2020 Mar 2. pii: 10.1038/s41587-020-0434-2. doi: 
            10.1038/s41587-020-0434-2."
COMMENT     Sequence Label: pv9_MBD1_MBD-R22A-13XL-BASU
FEATURES             Location/Qualifiers
     source          1..7413
                     /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"
     protein_bind    409..442
                     /label=loxP511
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are 
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     enhancer        447..826
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          1104..2120
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     2044..2066
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2128..2147
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2198..2207
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2207..2230
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2516..2536
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2822..2848
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    3726..4188
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3782..3799)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4009..4028
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4202..4801
                     /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(4202..4221)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4698..4718
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4970..5091
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5007..5026)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5061..5080
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(5125..5158)
                     /label=loxP511
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are 
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     primer_bind     complement(5192..5208)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5192..5208)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5205..5227)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5216..5232
                     /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(5240..5246,5247..5264,5265..5270))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5285..5306
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5423..5440)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5594..6182)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5674..5693)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6353..7144,7145..7213))
                     /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     6976..6995
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7214..7318)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7386..7404
                     /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 ctcgagccat aacttcgtat
      421 aatgtatact atacgaagtt atatcggaca ttgattattg actagttatt aatagtaatc
      481 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
      541 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
      601 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
      661 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
      721 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
      781 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc
      841 cacgttctgc ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt
      901 tattttttaa ttattttgtg cagcgatggg ggcggggggg gggggggsgc gcgccrgggg
      961 gggggggggg gggggggggg sggggsgggg sgrggcggag aggtgcggcg gcagccaatc
     1021 agagcggcgc gctccgaaag tttcctttta tggcgaggcg gcggcggcgg cggccctata
     1081 aaaagcgaag cgcgcggcgg gcgggagtcg ctgcgcgctg ccttcgcccc gtgccccgct
     1141 ccgccgccgc ctcgcgccgc ccgccccggc tctgactgac cgcgttactc ccacaggtga
     1201 gcgggcggga cggcccttct cctccgggct gtaattagcg cttggtttaa tgacggcttg
     1261 tttcttttct gtggctgcgt gaaagccttg aggggctccg ggagggccct ttgtgcgggg
     1321 ggagcggctc ggggggtgcg tgcgtgtgtg tgtgcgtggg gagcgccgcg tgcggctccg
     1381 cgctgcccgg cggctgtgag cgctgcgggc gcggcgcggg gctttgtgcg ctccgcagtg
     1441 tgcgcgaggg gagcgcggcc gggggcggtg ccccgcggtg cggggggggc tgcgagggga
     1501 acaaaggctg cgtgcggggt gtgtgcgtgg gggggtgagc agggggtgtg ggcgcgtcgg
     1561 tcgggctgca accccccctg cacccccctc cccgagttgc tgagcacggc ccggcttcgg
     1621 gtgcggggct ccgtacgggg cgtggcgcgg ggctcgccgt gccgggcggg gggtggcggc
     1681 aggtgggggt gccgggcggg gcggggccgc ctcgggccgg ggagggctcg ggggaggggc
     1741 gcggcggccc ccggagcgcc ggcggctgtc gaggcgcggc gagccgcagc cattgccttt
     1801 tatggtaatc gtgcgagagg gcgcagggac ttcctttgtc ccaaatctgt gcggagccga
     1861 aatctgggag gcgccgccgc accccctcta gcgggcgcgg ggcgaagcgg tgcggcgccg
     1921 gcaggaagga aatgggcggg gagggccttc gtgcgtcgcc gcgccgccgt ccccttctcc
     1981 ctctccagcc tcggggctgt ccgcgggggg acggctgcct tcggggggga cggggcaggg
     2041 cggggttcgg cttctggcgt gtgaccggcg gctctagagc ctctgctaac catgttcatg
     2101 ccttcttctt tttcctacag ctcctgggca acgtgctggt tattgtgctg tctcatcatt
     2161 ttggcaaaga attcattatt gtttttcctc ctttgccgcc accatggact acaaagacga
     2221 tgacgacaag gatgaagagt ctggttctgt ttctggtggt gatgctgagt cctggcagga
     2281 ctgcccagcc ctgggccctg gctggaaacg ccgagagtcc tttgcaaagt caggggcctc
     2341 gtttggacgc tcagacatct attaccagag ccccacagga gaaaagatcc gaagcaaagt
     2401 tgagctgact cggtacttgg gccctgcatg tgatcttacc ctgtttgact tcagacaagg
     2461 caccttatgc catcccatcg gtggcggagg ctctggaggt ggaggctccg caccccccaa
     2521 aaagaaacgc aaggttggat accgggatcc acacgtgggt aggagcggtt cgttcggatc
     2581 ccatcttttt tcggaattcg gatccggtgg aggcgggtct ggaggcgggg gtagtggcgg
     2641 gggtggaagc gggggtggag gcgggtcggg tggcggaggt agcggaggcg gtggaagtgg
     2701 aggcggtgga agtggtggag gaggttctgg tggcggaggt agcggtggcg gaggtagcgg
     2761 aggcggtgga agtggtggcg gaggtagcgg aggcggtgga tcggcgcgcc gtttcaagaa
     2821 ctacccatac gatgtgcctg actatgctgg caagttgtct gaatcagaga ttaggtttgg
     2881 actgaaaacg gaggtgatgg gccaacatct tatctatcac gatgttttgt ccagcacaca
     2941 aaagacagcc catgaattgg caaataacaa tgcccccgag ggtaccctcg tggttgcaga
     3001 taaacagaca gccggtaggg gcgggatgag ccgagtgtgg cattctcaag aaggaaacgg
     3061 agtatggatg agtttgatat tgcgcccgga cattccgctc caaaagacgc cccaacttac
     3121 cttgctggca gctgtagcgg tggttcaggg aatagaagag gccgcgggca tacagactga
     3181 tataaagtgg ccgaacgaca tactcattaa cgggaaaaaa acagtaggga tacttaccga
     3241 aatgcaagct gaggaggatc gagtgcgctc cgtgattatt ggtattggca ttaatgtaaa
     3301 tcagcaaccc aatgatttcc ccgatgagct caaggacatt gcgacgagtc tgtcccaggc
     3361 agctggggaa aagatcgaca gggcaggtgt aatccagcat atcttgcttt gtttcgaaaa
     3421 gcgataccga gactacatga cccacggttt cacgcccatc aagctcttgt gggagtcata
     3481 cgccctcggt attggcacga acatgagggc gagaactctc aacgggacat tttacggtaa
     3541 ggcgttgggt attgacgacg aaggggtact tttgttggaa actaatgaag gcattaagaa
     3601 gatatacagc gcggacatca gccttaggtg aaacgaggta gcggccgccg aagccgcttg
     3661 gaataaggcc ggtgtgcgtt tgtctatatg ttattttcca ccatattgcc gtcttttggc
     3721 aatgtgaggg cccggaaacc tggccctgtc ttcttgacga gcattcctag gggtctttcc
     3781 cctctcgcca aaggaatgca aggtctgttg aatgtcgtga aggaagcagt tcctctggaa
     3841 gcttcttgaa gacaaacaac gtctgtagcg accctttgca ggcagcggaa ccccccacct
     3901 ggcgacaggt gcctctgcgg ccaaaagcca cgtgtataag atacacctgc aaaggcggca
     3961 caaccccagt gccacgttgt gagttggata gttgtggaaa gagtcaaatg gctctcctca
     4021 agcgtattca acaaggggct gaaggatgcc cagaaggtac cccattgtat gggatctgat
     4081 ctggggcctc ggtgcacatg ctttacatgt gtttagtcga ggttaaaaaa acgtctaggc
     4141 cccccgaacc acggggacgt ggttttcctt tgaaaaacac gatgataata tggccacaac
     4201 catgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt
     4261 acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga
     4321 ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga
     4381 catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga
     4441 gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg
     4501 ttcccggctg gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga
     4561 gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg
     4621 cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct
     4681 ggagacctcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc
     4741 cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg
     4801 acgcccgccc cacgacccgc agcgcccgac cgaaaggagc gcagtgctac gagatttcga
     4861 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     4921 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     4981 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     5041 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     5101 tataccgtgg atccggccgg ccccataact tcgtatagta tacattatac gaagttatcc
     5161 cctgcaggca tgcaagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     5221 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     5281 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     5341 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     5401 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     5461 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     5521 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     5581 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     5641 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     5701 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     5761 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     5821 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     5881 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     5941 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     6001 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     6061 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     6121 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     6181 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     6241 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     6301 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     6361 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     6421 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     6481 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     6541 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     6601 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     6661 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     6721 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     6781 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     6841 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     6901 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     6961 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     7021 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     7081 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     7141 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     7201 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     7261 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     7321 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     7381 ataaaaatag gcgtatcacg aggccctttc gtc
//
