LOCUS       sequence-301992-        7195 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For transient expression of mouse BORIS (CTCFL) tagged with eGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7195)
  AUTHORS   Nora EP, Caccianini L, Fudenberg G, So K, Kameswaran V, Nagle A, 
            Uebersohn A, Hajj B, Saux AL, Coulon A, Mirny LA, Pollard KS, Dahan 
            M, Bruneau BG
  TITLE     Molecular basis of CTCF binding polarity in genome folding.
  JOURNAL   Nat Commun. 2020 Nov 5;11(1):5612. doi: 10.1038/s41467-020-19283-x.
  PUBMED    33154377
REFERENCE   2  (bases 1 to 7195)
  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 
            Commun."; date: "2020-11-5"; volume: "11(1):5612. doi"; pages: " 
            10.1038/s41467-020-19283-x"
COMMENT     Sequence Label: pKS031 - pCAGGS-3XFLAG-BORIS-eGFP-bpA
FEATURES             Location/Qualifiers
     source          1..7195
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1670
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1594..1616
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1678..1697
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1757..1822
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             join(3755..3757,3758..4468)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(3755..3757,3758..4468)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(3797..3818)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4058..4077)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4405..4426
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4556..4575)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4621..4676
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4675..4694)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5035..5051)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5035..5051)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5048..5070)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5059..5075
                     /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(5083..5089,5090..5107,5108..5113))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5128..5149
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5274..5291)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5445..6033)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5525..5544)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6204..6995,6996..7064))
                     /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     6827..6846
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7065..7169)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt
     1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg
     1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1681 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attcctcgag aattctcacg
     1741 cgtgaattcc accatggact ataaggacca cgacggagac tacaaggatc atgatattga
     1801 ttacaaagac gatgacgata agaccggtgc tgccgctgag gtccctgtcc cttctgggta
     1861 cttcacccag atcaaagagc agaagttgaa gcctggagac ctagaggagg agaaagagga
     1921 ggacggggta caaagagtgg aagcccagga gggagttgtc aaggaggtgg aggccgagaa
     1981 cagttgcctg cttctggagg ccagggcccc ggtggagagc gacaggcgga tcctgaccct
     2041 gcaaacggtg cacctggagt cccaggatgt gcacctacag gggctgggat ggctgagcgt
     2101 gccacactct gaggagcttt cagggacggt accagaggcg gaaggcatac tgcagttgcc
     2161 atccgtgctg tggctcgacc cagagcccca gctcagcctt cagcattgcg tgacggtcag
     2221 catcccggaa gagctgtacc caccagagga gctgcagcgg atacattttc acctgctgag
     2281 agagaatgtg ctaatggccg aggagaaccc agagttaaca ccagacttgg acgaaagcac
     2341 agccctgaaa aagcccgaag aagatgaaaa ggaccagctc ccgccccagg gagagacaga
     2401 caagagagaa gagaggttgc tccttctgga aatgaaacca aaagagggaa aagacgacga
     2461 aattgtcctg accatttccc atctaagcct cgaagaacag caagatccac cagcggccaa
     2521 tcagacaagt gtgccgggag ccaaagccgc aaaaccaaaa cggcggaggc agaccaaggg
     2581 aaagcctcag agctttcagt gtgacacctg cccgttcact tcctccaagc tctcaacttt
     2641 caatcgtcac atcaaaattc acagcaatga gaggccacac ctgtgtcacc tgtgcctgaa
     2701 ggccttccgg actgtcactc ttcttaggaa ccatgtgaac acccacacag gaaccaggcc
     2761 ccacaagtgc agggactgcg acatggcgtt tgtcaccagt ggagaactcg tccggcacag
     2821 gcgttacaaa cacacttatg agaagccctt caagtgctcc ctgtgcaagt acgccagcgt
     2881 cgaggcaagc aagatgaagc gtcacatccg ctcacacacg ggtgagcgtc ccttccagtg
     2941 ttgccagtgt gcttacgcca gcagggactc ctacaagctg aagcgccaca tgaggacaca
     3001 ctcaggtgag aagccgtatg aatgtcccac ctgtcacgtc cggttcaccc agagcgggac
     3061 catgaaaatc catatagcac agaagcacgg agagaatgtg cccaaatacg agtgtcccca
     3121 ctgtgccacc atcatcgcga ggaagagcga cctgcgtgtc catctgcgta acctgcacag
     3181 ccagagcccg gaggagatga agtgccgata ctgtcccgct ggcttccatg agcgctatgc
     3241 cctcattcag caccagagga cccacaagaa cgagaagaag ttcaagtgca agcagtgcga
     3301 ttacgcgtgc aagcaggagc gatgcttgaa ggcgcacatg cgcatgcaca caggagagaa
     3361 gcccttctcc tgcctggcct gcaacaagca cttccgacag aagcagctac tgaccgtgca
     3421 cctgaggaag taccatgacc cgaacttcgt ccccaatctg cacctgtgcc tcaagtgtga
     3481 taaacgtttc tcccgctgga gtaacctgca gagacacaga aagaagtgtg acccggagca
     3541 tgagacgtta gcccccaaca aggacaggag accagtgaca aggacacagg cctcggaggg
     3601 agaagcagga cacaaggaag gggagcctca gtgccctggg gagcaggctc tgggccacca
     3661 aggagaagca gcggggagcc agagcccaga ccacggcctt acctgcgaga taatctttaa
     3721 catgatggat aagcttaagg ggggagcagg aggcgtgagc aagggcgagg agctgttcac
     3781 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
     3841 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac
     3901 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca
     3961 gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     4021 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg
     4081 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
     4141 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
     4201 cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca
     4261 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
     4321 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa
     4381 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
     4441 cactctcggc atggacgagc tgtacaagtc gcgataagct agctcaggtg caggctgcct
     4501 atcagaaggt ggtggctggt gtggccaatg ccctggctca caaataccac tgagatcttt
     4561 ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg acttctggct
     4621 aataaaggaa atttattttc attgcaatag tgtgttggaa ttttttgtgt ctctcactcg
     4681 gaaggacata tgggagggca aatcatttaa aacatcagaa tgagtatttg gtttagagtt
     4741 tggcaacata tgccatatgc tggctgccat gaacaaaggt ggctataaag aggtcatcag
     4801 tatatgaaac agccccctgc tgtccattcc ttattccata gaaaagcctt gacttgaggt
     4861 tagatttttt ttatattttg ttttgtgtta tttttttctt taacatccct aaaattttcc
     4921 ttacatgttt tactagccag atttttcctc ctctcctgac tactcccagt catagctgtc
     4981 cctcttctct tatgaagatc cctcgacctg cagcccaagc ttggcgtaat catggtcata
     5041 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     5101 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     5161 ctcactgccc gctttccagt cgggaaacct gtcgtgccag cggatccgct gcattaatga
     5221 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     5281 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     5341 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     5401 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     5461 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     5521 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     5581 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcaa
     5641 tgctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     5701 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     5761 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     5821 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     5881 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     5941 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     6001 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     6061 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     6121 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     6181 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     6241 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     6301 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     6361 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     6421 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     6481 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     6541 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     6601 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     6661 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     6721 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     6781 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     6841 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     6901 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     6961 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     7021 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     7081 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     7141 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctg
//
