LOCUS       sequence-218256-        6774 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Reporter for Alt-EJ, variant of EJ7-GFP. Double strand breaks are 
            induced with the 7a.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6774)
  AUTHORS   Bhargava R, Sandhu M, Muk S, Lee G, Vaidehi N, Stark JM
  TITLE     C-NHEJ without indels is robust and requires synergistic function of
            distinct XLF domains.
  JOURNAL   Nat Commun. 2018 Jun 27;9(1):2484. doi: 10.1038/s41467-018-04867-5.
  PUBMED    29950655
REFERENCE   2  (bases 1 to 6774)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2018-06-27"; volume: "9(1):2484. doi"; pages: " 
            10.1038/s41467-018-04867-5"
COMMENT     Sequence Label: pCMV6-AC 4-uHOM
FEATURES             Location/Qualifiers
     source          1..6774
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     126..148
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     140..157
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     141..157
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        318..697
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        698..901
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     852..872
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     927..946
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        927..945
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      960..969
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1011..1032)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1322..1341)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1669..1690
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1867..1920
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     primer_bind     complement(1954..1970)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    1981..2603
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2180..2199)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2627..2647)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2632..2961
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2812..2947
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3028..3822
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3082..3101)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3692..3711
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3996..4117
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4033..4052)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4087..4106
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4166..4182)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4179..4201)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4190..4206
                     /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(4214..4220,4221..4238,4239..4244))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4259..4280
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4397..4414)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4568..5156)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4648..4667)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5327..6118,6119..6187))
                     /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     5950..5969
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6188..6292)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6319..6747
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6406..6425)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6616..6637
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
       61 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
      121 gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagctgatct atacattgaa
      181 tcaatattgg caattagcca tattagtcat tggttatata gcataaatca atattggcta
      241 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      301 aatatgaccg ccatgttgac attgattatt gactagttat taatagtaat caattacggg
      361 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      421 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      481 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      541 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      601 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      661 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      721 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      781 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      841 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      901 tcgtttagtg aaccgtcaga attttgtaat acgactcact atagggcggc cgggaattcg
      961 ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     1021 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     1081 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     1141 ccaccctcgt gaccaccctg acctacggct aaggaagata tctagggata acagggtaat
     1201 cccgggggcc tattacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca
     1261 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     1321 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     1381 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     1441 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     1501 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     1561 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     1621 gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     1681 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtaaggccg
     1741 gccagccacg gcttcccccc tgaggtggcc gctcaggacg atggcaccct gcccatgagc
     1801 tgcgcccagg agagcggcat ggacaggcac cccgccgctt gcgccagcgc taggatcaac
     1861 gtgggtgagg gcagaggaag tcttctaaca tgcggtgacg tggaggagaa tccgggccct
     1921 gtggtaccct cgaggtttaa acggccggcc gcggtcatag ctgtttcctg aacagatccc
     1981 gggtggcatc cctgtgaccc ctccccagtg cctctcctgg ccctggaagt tgccactcca
     2041 gtgcccacca gccttgtcct aataaaatta agttgcatca ttttgtctga ctaggtgtcc
     2101 ttctataata ttatggggtg gaggggggtg gtatggagca aggggcaagt tgggaagaca
     2161 acctgtaggg cctgcggggt ctattgggaa ccaagctgga gtgcagtggc acaatcttgg
     2221 ctcactgcaa tctccgcctc ctgggttcaa gcgattctcc tgcctcagcc tcccgagttg
     2281 ttgggattcc aggcatgcat gaccaggctc agctaatttt tgtttttttg gtagagacgg
     2341 ggtttcacca tattggccag gctggtctcc aactcctaat ctcaggtgat ctacccacct
     2401 tggcctccca aattgctggg attacaggcg tgaaccactg ctcccttccc tgtccttctg
     2461 attttaaaat aactatacca gcaggaggac gtccagacac agcataggct acctggccat
     2521 gcccaaccgg tgggacattt gagttgcttg cttggcactg tcctctcatg cgttgggtcc
     2581 actcagtaga tgcctgttga attgggtacg cggccagctt ggctgtggaa tgtgtgtcag
     2641 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     2701 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     2761 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     2821 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     2881 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     2941 ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat
     3001 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct
     3061 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     3121 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     3181 gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc
     3241 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     3301 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     3361 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     3421 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     3481 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     3541 gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc
     3601 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     3661 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     3721 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     3781 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg
     3841 aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct
     3901 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     3961 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg
     4021 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     4081 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct
     4141 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     4201 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     4261 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     4321 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     4381 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     4441 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     4501 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     4561 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     4621 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     4681 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     4741 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     4801 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     4861 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     4921 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     4981 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     5041 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     5101 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     5161 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     5221 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     5281 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     5341 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     5401 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     5461 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     5521 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     5581 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     5641 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     5701 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     5761 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     5821 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     5881 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     5941 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     6001 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     6061 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     6121 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     6181 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     6241 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     6301 cccgaaaagt gccacctgac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     6361 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     6421 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     6481 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     6541 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     6601 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     6661 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     6721 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca attt
//
