LOCUS       sequence-430977-        8084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Homologous recombination donor to insert halo tag at the N terminal 
            of the endogenous XLF locus..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8084)
  AUTHORS   Mikhova M, Goff NJ, Janovic T, Heyza JR, Meek K, Schmidt JC
  TITLE     Single-molecule imaging reveals the kinetics of non-homologous 
            end-joining in living cells.
  JOURNAL   Nat Commun. 2024 Nov 23;15(1):10159. doi: 
            10.1038/s41467-024-54545-y.
  PUBMED    39578493
REFERENCE   2  (bases 1 to 8084)
  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: "2024-11-23"; volume: "15(1):10159. doi"; pages: " 
            10.1038/s41467-024-54545-y"
COMMENT     Sequence Label: N-terminal Halo-XLF
FEATURES             Location/Qualifiers
     source          1..8084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     254..271
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  405..629
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(696..1229)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(1418..1423,1424..1440,1441..1446))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     polyA_signal    1954..2002
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     2027..2046
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     promoter        complement(2138..2247)
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     promoter        2271..2362
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     2287..2304
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             3074..3139
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     protein_bind    3140..3173
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     polyA_signal    3180..3228
                     /note="synthetic polyadenylation signal"
     CDS             complement(3229..3828)
                     /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(3312..3332)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     3809..3828
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(3835..4151)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3849..3984
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     protein_bind    4158..4191
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             4200..5090
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             5103..5123
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     polyA_signal    5468..5602
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5505..5524)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5559..5578
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  5631..5796
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      5980..6434
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6067..6086)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6276..6297
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6460..6564
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6565..6633,6634..7425)
                     /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     complement(6783..6802)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(7596..8084,1..100)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
       61 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
      121 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
      181 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
      241 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
      301 tctccttacg catctgtgcg gtatttcaca ccgcatagac cagccgcgta acctggcaaa
      361 atcggttacg gttgagtaat aaatggatgc cctgcgtaag cgggtgtggg cggacaataa
      421 agtcttaaac tgaacaaaat agatctaaac tatgacaata aagtcttaaa ctagacagaa
      481 tagttgtaaa ctgaaatcag tccagttatg ctgtgaaaaa gcatactgga cttttgttat
      541 ggctaaagca aactcttcat tttctgaagt gcaaattgcc cgtcgtatta aagaggggcg
      601 tggccaaggg catggtaaag actatattcg cggcgttgtg acaatttacc gaacaactcc
      661 gcggccggga agccgatctc ggcttgaacg aattgttagg tggcggtact tgggtcgata
      721 tcaaagtgca tcacttcttc ccgtatgccc aactttgtat agagagccac tgcgggatcg
      781 tcaccgtaat ctgcttgcac gtagatcaca taagcaccaa gcgcgttggc ctcatgcttg
      841 aggagattga tgagcgcggt ggcaatgccc tgcctccggt gctcgccgga gactgcgaga
      901 tcatagatat agatctcact acgcggctgc tcaaacttgg gcagaacgta agccgcgaga
      961 gcgccaacaa ccgcttcttg gtcgaaggca gcaagcgcga tgaatgtctt actacggagc
     1021 aagttcccga ggtaatcgga gtccggctga tgttgggagt aggtggctac gtctccgaac
     1081 tcacgaccga aaagatcaag agcagcccgc atggatttga cttggtcagg gccgagccta
     1141 catgtgcgaa tgatgcccat acttgagcca cctaactttg ttttagggcg actgccctgc
     1201 tgcgtaacat cgttgctgct gcgtaacatc gttgctgctc cataacatca aacatcgacc
     1261 cacggcgtaa cgcgcttgct gcttggatgc ccgaggcata gactgtacaa aaaaacagtc
     1321 ataacaagcc atgaaaaccg ccactgcgcc gttaccaccg ctgcgttcgg tcaaggttct
     1381 ggaccagttg cgtgagcgca tacgctactt gcattacagt ttacgaaccg aacaggctta
     1441 tgtcaactgg gttcgtgcct tcatccgttt ccacggtgtg cgtcacccgg caaccttggg
     1501 cagcagcgaa gtcgaggcat ttctgtcctg gctggcgaac gagcgcaagg tttcggtctc
     1561 cacgcatcgt caggcattgg cggccttgct gttcttctac ggcaaggtgc tgtgcacgga
     1621 tctgcccttg cttcaggaga tcggtagacc tcggccgtcg cggcgcttgc cggtggtgct
     1681 gaccccggat gaagtggttc gcatcctcgg ttttctggaa ggcgagcatc gtttgttcgc
     1741 ccaggactct agctatagtt ctagtggttg gctacgtacc cgtagtggct atggcagggc
     1801 ttgccgcccc gacgttggct gcgagccctg ggccttcacc cgaacttggg ggttggggtg
     1861 gggaaaagga agaaacgcgg gcgtattggt cccaatgggg tctcggtggg gtatcgacag
     1921 agtgccagcc ctgggaccga accccgcgtt tatgaacaaa cgacccaaca cccgtgcgtt
     1981 ttattctgtc tttttattgc cgtcatagcg cgggttcctt ccggtattgt ctccttccgt
     2041 gtttcagtta gcctccccca tctcccggta ccgcatgcta tgcatcagct gctagcacca
     2101 tggctcgaga tcccgggtga tcaagtcttc gtcgagtgat tgtaaataaa atgtaattta
     2161 cagtatagta ttttaattaa tatacaaatg atttgataat aattcttatt taactataat
     2221 atattgtgtt gggttgaatt aaaggtccgt atactccgga atattaatag atcatggaga
     2281 taattaaaat gataaccatc tcgcaaataa ataagtattt tactgttttc gtaacagttt
     2341 tgtaataaaa aaacctataa atattccgga ttattcatac cgtcccacca tcgggcgcgg
     2401 atcccggtcc gaagcgcgcg gaattcaaag gcctacgtcg acgagctcac tagtcgcggc
     2461 cgctttcgaa tctagagcct gcagtctcga caagcttgtc gagaagtact agaggatcat
     2521 aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc
     2581 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt tgacactatg acttatgatg
     2641 ttttataaga caagaactat tccttctctt ctagttctgg gaaaggatag gagtgacata
     2701 tttattagga atgtgataaa gatggaacaa gaagggttta ggtttccacg ttatgttata
     2761 gtattagaaa gcagaaatac taacagcaat atgctgaggt ttaaacacta tggaagaaag
     2821 attttctgtc ctctaggaat gtaatatggg aaaaaaaaaa ccctcaagat tttaaatata
     2881 cttttagtgg ccaggaggta gtaaagtcag tgctcagctg gttgtatact gggagaacgt
     2941 aaatccatgc cttctatttt atcagggagg attttcatga gattttagtt gatacagact
     3001 ggtttgggtg gtgccctccc taggccagca tgaccctgag gctctttact aaatgaggac
     3061 ttttttgcag atggactaca aagaccatga cggtgattat aaagatcatg atatcgatta
     3121 caaggatgac gatgacaaga taacttcgta taatgtatgc tatacgaagt tataagcttc
     3181 acacaaaaaa ccaacacaca gatgtaatga aaataaagat attttatttc aggcaccggg
     3241 cttgcgggtc atgcaccagg tgcgcggtcc ttcgggcacc tcgacgtcgg cggtgacggt
     3301 gaagccgagc cgctcgtaga aggggaggtt gcggggcgcg gaggtctcca ggaaggcggg
     3361 caccccggcg cgctcggccg cctccactcc ggggagcacg acggcgctgc ccagaccctt
     3421 gccctggtgg tcgggcgaga cgccgacggt ggccaggaac cacgcgggct ccttgggccg
     3481 gtgcggcgcc aggaggcctt ccatctgttg ctgcgcggcc agccgggaac cgctcaactc
     3541 ggccatgcgc gggccgatct cggcgaacac cgcccccgct tcgacgctct ccggcgtggt
     3601 ccagaccgcc accgcggcgc cgtcgtccgc gacccacacc ttgccgatgt cgagcccgac
     3661 gcgcgtgagg aagagttctt gcagctcggt gacccgctcg atgtggcggt ccggatcgac
     3721 ggtgtggcgc gtggcggggt agtcggcgaa cgcggcggcg agggtgcgta cggccctggg
     3781 gacgtcgtcg cgggtggcga ggcgcaccgt gggcttgtac tcggtcatga gctttttgca
     3841 aaagcctagg cctccaaaaa agcctcctca ctacttctgg aatagctcag aggccgaggc
     3901 ggcctcggcc tctgcataaa taaaaaaaat tagtcagcca tggggcggag aatgggcgga
     3961 actgggcgga gttaggggcg ggatgggcgg agttaggggc gggactatgg ttgctgacta
     4021 attgagatgc atgctttgca tacttctgcc tgctggggag cctggggact ttccacacct
     4081 ggttgctgac taattgagat gcatgctttg catacttctg cctgctgggg agcctgggga
     4141 ctttccacac cggatccata acttcgtata atgtatgcta tacgaagtta tccggtacca
     4201 tggcagaaat cggtactggc tttccattcg acccccatta tgtggaagtc ctgggcgagc
     4261 gcatgcacta cgtcgatgtt ggtccgcgcg atggcacccc tgtgctgttc ctgcacggta
     4321 acccgacctc ctcctacgtg tggcgcaaca tcatcccgca tgttgcaccg acccatcgct
     4381 gcattgctcc agacctgatc ggtatgggca aatccgacaa accagacctg ggttatttct
     4441 tcgacgacca cgtccgcttc atggatgcct tcatcgaagc cctgggtctg gaagaggtcg
     4501 tcctggtcat tcacgactgg ggctccgctc tgggtttcca ctgggccaag cgcaatccag
     4561 agcgcgtcaa aggtattgca tttatggagt tcatccgccc tatcccgacc tgggacgaat
     4621 ggccagaatt tgcccgcgag accttccagg ccttccgcac caccgacgtc ggccgcaagc
     4681 tgatcatcga tcagaacgtt tttatcgagg gtacgctgcc gatgggtgtc gtccgcccgc
     4741 tgactgaagt cgagatggac cattaccgcg agccgttcct gaatcctgtt gaccgcgagc
     4801 cactgtggcg cttcccaaac gagctgccaa tcgccggtga gccagcgaac atcgtcgcgc
     4861 tggtcgaaga atacatggac tggctgcacc agtcccctgt cccgaagctg ctgttctggg
     4921 gcaccccagg cgttctgatc ccaccggccg aagccgctcg cctggccaaa agcctgccta
     4981 actgcaaggc tgtggacatc ggcccgggtc tgaatctgct gcaagaagac aacccggacc
     5041 tgatcggcag cgagatcgcg cgctggctgt cgacgctcga gatttccggc gagccaacca
     5101 ctgaggatct gtactttcag agcgataacg cgatcgcttc cggtaccgaa gaactcgagc
     5161 aaggcctgtt gatgcagcca tgggcgtggc tacagcttgc agagaactcc ctcttggcca
     5221 aggtttttat caccaagcag ggctatgcct tgttggtttc agatcttcaa caggtgtggc
     5281 atgaacaggt ggacactagt gtggtcagcc agcgagccaa ggtaagtatg aggagaagta
     5341 ctgctgtgtc ggggatgtga acatcaacca gcttgctggc ctgtagtttc ccaaggaagg
     5401 ttgaatcggg ttgagacaat tccagttaat gtcctgtact tctacctgta cttctgcctg
     5461 gaatcttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     5521 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     5581 tgtatcttat catgtctgga tctgatcact gcttgagcct aggagatccg aaccagataa
     5641 gtgaaatcta gttccaaact attttgtcat ttttaatttt cgtattagct tacgacgcta
     5701 cacccagttc ccatctattt tgtcactctt ccctaaataa tccttaaaaa ctccatttcc
     5761 acccctccca gttcccaact attttgtccg cccacagcgg ggcatttttc ttcctgttat
     5821 gtttttaatc aaacatcctg ccaactccat gtgacaaacc gtcatcttcg gctacttttt
     5881 ctctgtcaca gaatgaaaat ttttctgtca tctcttcgtt attaatgttt gtaattgact
     5941 gaatatcaac gcttatttgc agcctgaatg gcgaatggga cgcgccctgt agcggcgcat
     6001 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
     6061 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     6121 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     6181 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
     6241 ttcgcccttt gacgttggag tccacgttct taatagtgga ctcttgttcc aaactggaac
     6301 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     6361 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     6421 aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     6481 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     6541 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     6601 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     6661 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     6721 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     6781 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     6841 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     6901 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     6961 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     7021 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     7081 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     7141 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     7201 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     7261 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     7321 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     7381 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     7441 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     7501 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     7561 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     7621 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     7681 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     7741 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     7801 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     7861 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     7921 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     7981 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     8041 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagg
//
