LOCUS       sequence-437547-        3920 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To generate substrates to monitor Ku overloading and its impact on 
            transcription at the DNA ends vicinity..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3920)
  AUTHORS   Bossaert M, Moreno A, Peixoto A, Pillaire MJ, Chanut P, Frit P, 
            Calsou P, Loparo JJ, Britton S
  TITLE     Identification of the main barriers to Ku accumulation in chromatin.
  JOURNAL   bioRxiv [Preprint]. 2024 Jan 4:2024.01.03.574002. doi: 
            10.1101/2024.01.03.574002.
  PUBMED    38260538
REFERENCE   2  (bases 1 to 3920)
  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: "bioRxiv 
            [Preprint]. 2024 Jan 4:2024.01.03.574002. doi: 
            10.1101/2024.01.03.574002."
COMMENT     Sequence Label: pUC18-GFP
FEATURES             Location/Qualifiers
     source          1..3920
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     108..126
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(164..186)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     286..305
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     499..521
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     513..530
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     514..530
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    570..794
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     CDS             complement(join(795..1508,1509..1511,1512..1514))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(798..1508,1509..1511,1512..1514))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(840..861)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1189..1208
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1448..1469
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1511..1520
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(1553..1749)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        complement(1561..1771)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1567..1702
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1621..1640)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     1795..1815
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     primer_bind     complement(1834..1850)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1834..1850)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1847..1869)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1858..1874
                     /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(1882..1888,1889..1906,1907..1912))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1927..1948
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2065..2082)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2236..2824)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2316..2335)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2995..3786,3787..3855))
                     /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     3618..3637
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(3856..3920,1..40))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
       61 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
      121 cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc
      181 tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg
      241 gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga
      301 ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat
      361 accgcatcag gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc
      421 gggcctcttc gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt
      481 gggtaacgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt gccaagcttg
      541 catgcctgca ggtcgactct agaggatccc catagagccc accgcatccc cagcatgcct
      601 gctattgtct tcccaatcct cccccttgct gtcctgcccc accccacccc ccagaataga
      661 atgacaccta ctcagacaat gcgatgcaat ttcctcattt tattaggaaa ggacagtggg
      721 agtggcacct tccagggtca aggaaggcac gggggagggg caaacaacag atggctggca
      781 actagaaggc acagttactt gtacagctcg tccatgccga gagtgatccc ggcggcggtc
      841 acgaactcca gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct cagggcggac
      901 tgggtgctca ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat gggggtgttc
      961 tgctggtagt ggtcggcgag ctgcacgctg ccgtcctcga tgttgtggcg gatcttgaag
     1021 ttcaccttga tgccgttctt ctgcttgtcg gccatgatat agacgttgtg gctgttgtag
     1081 ttgtactcca gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat gcccttcagc
     1141 tcgatgcggt tcaccagggt gtcgccctcg aacttcacct cggcgcgggt cttgtagttg
     1201 ccgtcgtcct tgaagaagat ggtgcgctcc tggacgtagc cttcgggcat ggcggacttg
     1261 aagaagtcgt gctgcttcat gtggtcgggg tagcggctga agcactgcac gccgtaggtc
     1321 agggtggtca cgagggtggg ccagggcacg ggcagcttgc cggtggtgca gatgaacttc
     1381 agggtcagct tgccgtaggt ggcatcgccc tcgccctcgc cggacacgct gaacttgtgg
     1441 ccgtttacgt cgccgtccag ctcgaccagg atgggcacca ccccggtgaa cagctcctcg
     1501 cccttgctca ccatggtggc gcgaaaatgg atatacaagc tcccgggagc tttttgcaaa
     1561 agcctaggcc tccaaaaaag cctcctcact acttctggaa tagctcagag gcagaggcgg
     1621 cctcggcctc tgcataaata aaaaaaatta gtcagccatg gggcggagaa tgggcggaac
     1681 tgggcggagt taggggcggg atgggcggag ttaggggcgg gactatggtt gctgactaat
     1741 tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt ccacacccta
     1801 actgacacac attccacagg aattcgtaat catgtcatag ctgtttcctg tgtgaaattg
     1861 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     1921 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     1981 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     2041 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     2101 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     2161 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     2221 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     2281 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     2341 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     2401 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     2461 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     2521 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     2581 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     2641 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     2701 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     2761 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     2821 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     2881 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     2941 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     3001 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     3061 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     3121 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     3181 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     3241 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     3301 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     3361 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     3421 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     3481 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     3541 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     3601 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     3661 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     3721 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     3781 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     3841 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     3901 tctcatgagc ggatacatat
//
