LOCUS       sequence-181118-        6912 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for expression of GFP-FLAG tagged human Ku80 (N-terminal 
            tag). Confers resistance to G418..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6912)
  AUTHORS   Britton S, Coates J, Jackson SP
  TITLE     A new method for high-resolution imaging of Ku foci to decipher 
            mechanisms of DNA double-strand break repair.
  JOURNAL   J Cell Biol. 2013 Jul 29.
  PUBMED    23897892
REFERENCE   2  (bases 1 to 6912)
  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: "J Cell 
            Biol. 2013 Jul 29."
COMMENT     Sequence Label: pEGFP-C1-FLAG-Ku80
FEATURES             Location/Qualifiers
     source          1..6912
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        52..355
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        356..559
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     510..530
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      598..607
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(604..606,607..609,610..1320)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(604..606,607..609,610..1320)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(649..670)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(910..929)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1257..1278
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1336..1359
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    3691..3812
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3728..3747)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3782..3801
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3819..4274)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3956..3977)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4168..4187
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4301..4405
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4407..4764
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4407..4598
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4430..4450)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4615..4750
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4799..5593
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4799..5593
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4853..4872)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5463..5482
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5781..5800)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5825..5872
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6201..6789
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6690..6709
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atagtaatca attacggggt cattagttca tagcccatat atggagttcc gcgttacata
       61 acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat
      121 aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga
      181 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc
      241 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt
      301 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat
      361 gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag
      421 tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc
      481 aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga
      541 ggtctatata agcagagctg gtttagtgaa ccgtcagatc cgctagcgct accggtcgcc
      601 accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
      661 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
      721 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
      781 accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg
      841 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
      901 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
      961 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     1021 cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga caagcagaag
     1081 aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc
     1141 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     1201 cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg
     1261 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
     1321 tccggactca gatctgatta caaggatgac gacgataagc tcgaggtgcg gtcggggaat
     1381 aaggcagctg ttgtgctgtg tatggacgtg ggctttacca tgagtaactc cattcctggt
     1441 atagaatccc catttgaaca agcaaagaag gtgataacca tgtttgtaca gcgacaggtg
     1501 tttgctgaga acaaggatga gattgcttta gtcctgtttg gtacagatgg cactgacaat
     1561 cccctttctg gtggggatca gtatcagaac atcacagtgc acagacatct gatgctacca
     1621 gattttgatt tgctggagga cattgaaagc aaaatccaac caggttctca acaggctgac
     1681 ttcctggatg cactaatcgt gagcatggat gtgattcaac atgaaacaat aggaaagaag
     1741 tttgagaaga ggcatattga aatattcact gacctcagca gccgattcag caaaagtcag
     1801 ctggatatta taattcatag cttgaagaaa tgtggcatct ccctgcaatt cttcttgcct
     1861 ttctcacttg gcaaggaaga tggaagtggg gacagaggag atggcccctt tcgcttaggt
     1921 ggccatgggc cttcctttcc actaaaagga attaccgaac agcaaaaaga aggtcttgag
     1981 atagtgaaaa tggtgatgat atctttagaa ggtgaagatg ggttggatga aatttattca
     2041 ttcagtgaga gtctgagaaa actgtgcgtc ttcaagaaaa ttgagaggca ttccattcac
     2101 tggccctgcc gactgaccat tggctccaat ttgtctataa ggattgcagc ctataaatcg
     2161 attctacagg agagagttaa aaagacttgg acagttgtgg atgcaaaaac cctaaaaaaa
     2221 gaagatatac aaaaagaaac agtttattgc ttaaatgatg atgatgaaac tgaagtttta
     2281 aaagaggata ttattcaagg gttccgctat ggaagtgata tagttccttt ctctaaagtg
     2341 gatgaggaac aaatgaaata taaatcggag gggaagtgct tctctgtttt gggattttgt
     2401 aaatcttctc aggttcagag aagattcttc atgggaaatc aagttctaaa ggtctttgca
     2461 gcaagagatg atgaggcagc tgcagttgca ctttcctccc tgattcatgc tttggatgac
     2521 ttagacatgg tggccatagt tcgatacgct tatgacaaaa gagctaatcc tcaagtcggc
     2581 gtggcttttc ctcatatcaa gcataactat gagtgtttag tgtatgtgca gctgcctttc
     2641 atggaagact tgcggcaata catgttttca tccttgaaaa acagtaagaa atatgctccc
     2701 accgaggcac agttgaatgc tgttgatgct ttgattgact ccatgagctt ggcaaagaaa
     2761 gatgagaaga cagacaccct tgaagacttg tttccaacca ccaaaatccc aaatcctcga
     2821 tttcagagat tatttcagtg tctgctgcac agagctttac atccccggga gcctctaccc
     2881 ccaattcagc agcatatttg gaatatgctg aatcctcccg ctgaggtgac aacgaaaagt
     2941 cagattcctc tctctaaaat aaagaccctt tttcctctga ttgaagccaa gaaaaaggat
     3001 caagtgactg ctcaggaaat tttccaagac aaccatgaag atggacctac agctaaaaaa
     3061 ttaaagactg agcaaggggg agcccacttc agcgtctcca gtctggctga aggcagtgtc
     3121 acctctgttg gaagtgtgaa tcctgctgaa aacttccgtg ttctagtgaa acagaagaag
     3181 gccagctttg aggaagcgag taaccagctc ataaatcaca tcgaacagtt tttggatact
     3241 aatgaaacac cgtattttat gaagagcata gactgcatcc gagccttccg ggaagaagcc
     3301 attaagtttt cagaagagca gcgctttaac aacttcctga aagcccttca agagaaagtg
     3361 gagattaaac aattaaatca tttctgggaa attgttgtcc aggatggaat tactctgatc
     3421 accaaagagg aagcctctgg aagttctgtc acagctgagg aagccaaaaa gtttctggcc
     3481 cccaaagaca aaccaagtgg agacacagca gctgtatttg aagaaggtgg tgatgtggac
     3541 gatttattgg acatgatata aggatccacc ggatctagat aactgatcat aatcagccat
     3601 accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg
     3661 aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac
     3721 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     3781 tgtggtttgt ccaaactcat caatgtatct taacgcgtaa attgtaagcg ttaatatttt
     3841 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat
     3901 cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt
     3961 ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt
     4021 ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag
     4081 gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg
     4141 aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc
     4201 gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc
     4261 gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     4321 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     4381 tcaataatat tgaaaaagga agagtcctga ggcggaaaga accagctgtg gaatgtgtgt
     4441 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     4501 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     4561 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     4621 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     4681 tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     4741 tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat cgtttcgcat
     4801 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg
     4861 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc
     4921 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca
     4981 agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct
     5041 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga
     5101 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg
     5161 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat
     5221 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga
     5281 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg
     5341 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg
     5401 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat
     5461 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct
     5521 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga
     5581 cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg
     5641 ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     5701 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     5761 caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg aacccgcgct
     5821 atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc
     5881 ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca ttggggccaa
     5941 tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga aggcccaggg
     6001 ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc atatatactt
     6061 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     6121 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     6181 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     6241 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     6301 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     6361 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     6421 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     6481 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     6541 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     6601 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     6661 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     6721 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     6781 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     6841 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccatg
     6901 cattagttat ta
//
