LOCUS       sequence-64913-e        4365 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 4365)
  AUTHORS   Monnat RJ Jr, Hackmann AF, Cantrell MA
  TITLE     Generation of highly site-specific DNA double-strand breaks in human
            cells by the homing endonucleases I-PpoI and I-CreI.
  JOURNAL   Biochem Biophys Res Commun. 1999 Feb 5;255(1):88-93.
  PUBMED    10082660
REFERENCE   2  (bases 1 to 4365)
  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: "Biochem 
            Biophys Res Commun."; date: "1999-02-5"; volume: "255(1)"; pages: 
            "88-93"
COMMENT     Sequence Label: pCNCre3
FEATURES             Location/Qualifiers
     source          1..4365
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        27..330
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        331..534
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     485..505
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     531..555
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          672..768
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     CDS             838..858
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    1504..1638
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1528..1547)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1582..1601
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1749..1765)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1749..1765)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1762..1784)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1773..1789
                     /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(1797..1803,1804..1821,1822..1827))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1842..1863
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1980..1997)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2151..2739)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2231..2250)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2910..3701,3702..3770))
                     /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     3533..3552
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3771..3875)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3943..3961
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3999..4021)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4121..4140
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4334..4356
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4348..4365
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4349..4365
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gaattcgagc ttgcatgcct gcaggtcgtt acataactta cggtaaatgg cccgcctggc
       61 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      121 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
      181 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
      241 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
      301 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
      361 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
      421 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
      481 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta
      541 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac
      601 cgggaccgat ccagcctccg gactctagag gatccggtac tcgaggaact gaaaaaccag
      661 aaagttaact ggtaagttta gtctttttgt cttttatttc aggtcccgga tccggtggtg
      721 gtgcaaatca aagaactgct cctcagtgga tgttgccttt acttctaggc ctgtacggaa
      781 gtgttacttc tgctctaaaa gctgcggaat tgtacccgcg gccaattcac catggctcca
      841 aaaaaaaagc gcaaagtggc gctgaataca aaatataata aagagttctt actctactta
      901 gcagggtttg tagacggtga cggtagcata atcgctcaaa ttaagcctaa tcagtcttat
      961 aaatttaagc atcagctatc actcacgttc caagtcacgc aaaagacaca gagacgttgg
     1021 tttttaggca aattagtgga tgaaattggg gttggttatg taagagatag gggtagcgtt
     1081 tcggattata ttctaagcga aatcaagcct ttgcataatt ttttaacaca actacaacct
     1141 tttctaaaac taaaacaaaa acaagcaaat ttagttttaa aaattattga acaacttccg
     1201 tcagcaaaag aatccccgga caaattctta gaagtttgta catgggtgga tcaaattgca
     1261 gctctgaatg attcgaagac gcgtaaaaca acttctgaaa ccgttcgtgc tgtgctagac
     1321 agtttaagtg aaaaaaagaa atcgtccccg tagagacttt ataaatttag ccaatctcta
     1381 aaagaatgtt tacatacaat ttatttattg ttgctcgatt tataggatat tttctcgaga
     1441 gtgggaaagt ataatacgcc gactcctggg atccccgggt accgagctcg aattggccgc
     1501 ggggatccag acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag
     1561 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata
     1621 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg
     1681 gaggtgtggg aggttttttc ggatcctcta gagtcgacct gcaggcatgc aagcttggcg
     1741 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     1801 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     1861 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     1921 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     1981 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     2041 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     2101 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     2161 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     2221 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     2281 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     2341 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     2401 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     2461 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     2521 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     2581 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     2641 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     2701 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     2761 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     2821 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     2881 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     2941 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3001 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3061 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3121 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     3181 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     3241 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     3301 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     3361 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     3421 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     3481 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     3541 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     3601 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     3661 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     3721 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     3781 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     3841 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     3901 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
     3961 ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
     4021 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
     4081 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
     4141 ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc
     4201 atcaggcgcc attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc
     4261 tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta
     4321 acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagt
//
