LOCUS       sequence-46758-e        5911 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5911)
  AUTHORS   Perez-Pinera P, Ousterout DG, Brown MT, Gersbach CA
  TITLE     Gene targeting to the ROSA26 locus directed by engineered zinc 
            finger nucleases.
  JOURNAL   Nucleic Acids Res. 2012 Apr 1;40(8):3741-52. Epub 2011 Dec 14.
  PUBMED    22169954
REFERENCE   2  (bases 1 to 5911)
  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: "Nucleic 
            Acids Res."; date: "2012-04-1"; volume: "40(8)"; pages: "3741-52. 
            Epub 2011 Dec 14"
COMMENT     Sequence Label: pCMV-RosaL6 ELD mutations
FEATURES             Location/Qualifiers
     source          1..5911
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        87..390
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        391..594
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     545..565
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     591..615
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        825..843
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     825..842
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     856..875
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             922..948
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             976..996
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      2159..2168
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2172..2306
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2209..2228)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2263..2282
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2375..2732
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2375..2566
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2398..2418)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2583..2718
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2752..2769)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2752..2768)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2761..2783)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2981..3436
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3068..3087)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3278..3299
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3453..3472)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3607..3629
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3682..3700)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3768..3872
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4058..4436
                     /label=ISS
                     /note="immunostimulatory sequence from the AmpR gene; 
                     contains unmethylated CpG dinucleotides in the context of 
                     5'-AACGTT-3' (Sato et al., 1996)"
     primer_bind     complement(4092..4111)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4905..5493
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5394..5413
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5647..5664
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(5817..5822,5823..5840,5841..5847)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5855..5871
                     /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)."
     primer_bind     5860..5882
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5879..5895
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5879..5895
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgaattga ttattgacag ttattaatag taatcattac ggggtcatta
       61 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
      121 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      181 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
      241 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
      301 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
      361 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
      421 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
      481 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
      541 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta
      601 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac
      661 cgggaccgat ccagcctccg cggccgggaa cggtgcattg gaacgcggat tccccgtgcc
      721 aagagtgacg taagtaccgc ctatagagtc tataggccca cccccttggc ttcgttagaa
      781 cgcggctaca attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat
      841 agaataacat ccactttgcc tttctctcca caggtgtcca ctcccaggtc caactgcacc
      901 tcggttctat cgattgacat gtacccatac gatgtcccag actacgcgaa ttcccctggg
      961 atcccaggca tggggcccaa aaagaaacgc aaagttgggc gcctcgagcc cggggaaaaa
     1021 ccctataaat gccccgagtg tggtaagtca ttctctcaaa gcggggattt aagaagacac
     1081 cagagaaccc acaccgggga aaagccctac aagtgtcctg agtgcggaaa gtctttctcc
     1141 actagcggtt cattagtaag acaccagagg acacacaccg gggagaaacc ttataaatgc
     1201 ccagaatgcg ggaaatcgtt cagtcaaaga gcacatttag aaagacatca acggacccac
     1261 accggggaaa aaccatacaa gtgccccgag tgtggcaaga gctttagtac ccacctcgac
     1321 ctgattagac accagcgcac ccacaccggg gaaaagccat ataagtgccc tgagtgtgga
     1381 aagagcttca gtaggaagga taaccttaaa aaccaccaaa gaacccacac cggggaaaag
     1441 ccctataagt gcccagagtg cgggaaatca ttctcacagc tggcacatct tagagcccac
     1501 cagcggaccc acaccggtgc ggccgcccgc gccctggtga agagcgagct ggaggagaag
     1561 aagtccgagc tgcggcacaa gctgaagtac gtgccccacg agtacatcga gctgatcgag
     1621 atcgccagga accccaccca ggaccgcatc ctggagatga aggtgatgga gttcttcatg
     1681 aaggtgtacg gctacagggg agagcacctg ggcggaagca gaaagcctga cggcgccatc
     1741 tatacagtgg gcagccccat cgattacggc gtgatcgtgg acacaaaggc ctacagcggc
     1801 ggctacaatc tgcctatcgg ccaggccgac gagatggaga gatacgtgga ggagaaccag
     1861 acacgggata agcacctcaa ccccaacgag tggtggaagg tgtaccctag cagcgtgacc
     1921 gagttcaagt tcctgttcgt gagcggccac ttcaagggca actacaaggc ccagctgacc
     1981 aggctgaacc acatcaccaa ctgcaatggc gccgtgctga gcgtggagga gctgctgatc
     2041 ggcggcgaga tgatcaaagc cggcaccctg acactggagg aggtgcggcg caagttcaac
     2101 aacggcgaga tcaacttctg attaattaac taatctagag tcgacctgca gaagcttggc
     2161 cgccatggcc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     2221 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     2281 tcaatgtatc ttatcatgtc tggatcggga attaattcgg cgcagcacca tggcctgaaa
     2341 taacctctga aagaggaact tggttaggta ccttctgagg cggaaagaac cagctgtgga
     2401 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     2461 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     2521 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     2581 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     2641 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     2701 gaggcttttt tggaggccta ggcatttgca aaaagctgtt aacagcttgg cactggccgt
     2761 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
     2821 acatcccccc ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     2881 acagttgcgt agcctgaatg gcgaatggcg cctgatgcgg tattttctcc ttacgcatct
     2941 gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca
     3001 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     3061 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     3121 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     3181 cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     3241 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     3301 acaacactca accctatctc gggctattct tttgatttat aagggatttt gccgatttcg
     3361 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     3421 ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3481 agccaactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
     3541 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     3601 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
     3661 agtattcttg aagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     3721 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     3781 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     3841 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     3901 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     3961 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     4021 acagcggtaa gatcctttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     4081 tttaaagttc tgctatgtgg cgcggtatta tcccgtgatg acgccgggca agagcaactc
     4141 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     4201 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     4261 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     4321 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     4381 gccataccaa acgacgagcg tgacaccacg atgccagcag caatggcaac aacgttgcgc
     4441 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     4501 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     4561 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     4621 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     4681 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     4741 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     4801 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     4861 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttcaga tccttttttt
     4921 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     4981 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     5041 ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     5101 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     5161 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     5221 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     5281 tacctacagc gtgagcattg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     5341 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     5401 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     5461 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     5521 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     5581 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     5641 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc
     5701 cccgcgcgtt ggccgattca ttaatccagc tggcacgaca ggtttcccga ctggaaagcg
     5761 ggcagtgagc gcaacgcaat taatgtgagt tacctcactc attaggcacc ccaggcttta
     5821 cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca
     5881 ggaaacagct atgaccatga ttacgaatta a
//
