LOCUS       sequence-373720-        7683 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cell line generation via recombination-mediated cassette exchange 
            (RMCE) and stable expression of CBX1.2xChromo-13XL-BASU.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7683)
  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 7683)
  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_CBX1_2xChromo-13XL-BASU
FEATURES             Location/Qualifiers
     source          1..7683
                     /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             2498..2518
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2786..2806
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3092..3118
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    3996..4458
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4052..4069)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4279..4298
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4472..5071
                     /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(4472..4491)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4968..4988
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5240..5361
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5277..5296)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5331..5350
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(5395..5428)
                     /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(5462..5478)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5462..5478)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5475..5497)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5486..5502
                     /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(5510..5516,5517..5534,5535..5540))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5555..5576
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5693..5710)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5864..6452)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5944..5963)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6623..7414,7415..7483))
                     /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     7246..7265
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7484..7588)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7656..7674
                     /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 gggggggggc gcgccrgggg
      961 gggggggggg gggggggggg gggggggggg sgrggsggrg rggkgcggcg 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 ggrgggctcg ggggrggggc
     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 gatgaagagg aatatgtggt
     2281 ggaaaaagtt cttgatcggc gagttgtcaa gggcaaggtg gaatatcttc taaagtggaa
     2341 gggtttctca gatgaggaca acacttggga gccagaagag aatctggatt gccctgacct
     2401 tattgctgag tttctacagt cacagaaaac agctcatgag acagataagt cagagggaat
     2461 cggtggcgga ggctctggag gtggaggctc cgcacccccc aaaaagaaac gcaaggttgg
     2521 ataccgggat ccagaagagg aatatgtggt ggaaaaagtt cttgatcggc gagttgtcaa
     2581 gggcaaggtg gaatatcttc taaagtggaa gggtttctca gatgaggaca acacttggga
     2641 gccagaagag aatctggatt gccctgacct tattgctgag tttctacagt cacagaaaac
     2701 agctcatgag acagataagt cagagggacc ggtcgccacc ttgagcatcg gtggcggagg
     2761 ctctggaggt ggaggctccg caccccccaa aaagaaacgc aaggttggat accgggatcc
     2821 acacgtgggt aggagcggtt cgttcggatc ccatcttttt tcggaattcg gatccggtgg
     2881 aggcgggtct ggaggcgggg gtagtggcgg gggtggaagc gggggtggag gcgggtcggg
     2941 tggcggaggt agcggaggcg gtggaagtgg aggcggtgga agtggtggag gaggttctgg
     3001 tggcggaggt agcggtggcg gaggtagcgg aggcggtgga agtggtggcg gaggtagcgg
     3061 aggcggtgga tcggcgcgcc gtttcaagaa ctacccatac gatgtgcctg actatgctgg
     3121 caagttgtct gaatcagaga ttaggtttgg actgaaaacg gaggtgatgg gccaacatct
     3181 tatctatcac gatgttttgt ccagcacaca aaagacagcc catgaattgg caaataacaa
     3241 tgcccccgag ggtaccctcg tggttgcaga taaacagaca gccggtaggg gcgggatgag
     3301 ccgagtgtgg cattctcaag aaggaaacgg agtatggatg agtttgatat tgcgcccgga
     3361 cattccgctc caaaagacgc cccaacttac cttgctggca gctgtagcgg tggttcaggg
     3421 aatagaagag gccgcgggca tacagactga tataaagtgg ccgaacgaca tactcattaa
     3481 cgggaaaaaa acagtaggga tacttaccga aatgcaagct gaggaggatc gagtgcgctc
     3541 cgtgattatt ggtattggca ttaatgtaaa tcagcaaccc aatgatttcc ccgatgagct
     3601 caaggacatt gcgacgagtc tgtcccaggc agctggggaa aagatcgaca gggcaggtgt
     3661 aatccagcat atcttgcttt gtttcgaaaa gcgataccga gactacatga cccacggttt
     3721 cacgcccatc aagctcttgt gggagtcata cgccctcggt attggcacga acatgagggc
     3781 gagaactctc aacgggacat tttacggtaa ggcgttgggt attgacgacg aaggggtact
     3841 tttgttggaa actaatgaag gcattaagaa gatatacagc gcggacatca gccttaggtg
     3901 aaacgaggta gcggccgccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg
     3961 ttattttcca ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc
     4021 ttcttgacga gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg
     4081 aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg
     4141 accctttgca ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca
     4201 cgtgtataag atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata
     4261 gttgtggaaa gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc
     4321 cagaaggtac cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt
     4381 gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt
     4441 tgaaaaacac gatgataata tggccacaac catgaccgag tacaagccca cggtgcgcct
     4501 cgccacccgc gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta
     4561 ccccgccacg cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca
     4621 agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg
     4681 cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga
     4741 gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga
     4801 aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt
     4861 ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc
     4921 ggccgagcgc gccggggtgc ccgccttcct ggagacctcc gcgccccgca acctcccctt
     4981 ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac
     5041 ctggtgcatg acccgcaagc ccggtgcctg acgcccgccc cacgacccgc agcgcccgac
     5101 cgaaaggagc gcagtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     5161 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     5221 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     5281 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     5341 caaactcatc aatgtatctt atcatgtctg tataccgtgg atccggccgg ccccataact
     5401 tcgtatagta tacattatac gaagttatcc cctgcaggca tgcaagcttg gcgtaatcat
     5461 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     5521 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     5581 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     5641 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     5701 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     5761 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     5821 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     5881 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     5941 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     6001 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     6061 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     6121 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     6181 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     6241 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     6301 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     6361 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     6421 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     6481 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     6541 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     6601 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     6661 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     6721 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     6781 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     6841 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     6901 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     6961 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     7021 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     7081 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     7141 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     7201 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     7261 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     7321 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     7381 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     7441 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     7501 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     7561 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct
     7621 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     7681 gtc
//
