LOCUS       sequence-14102-e        5416 bp    DNA     circular SYN 15-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5416)
  AUTHORS   Werdien D, Peiler G, Ryffel GU
  TITLE     FLP and Cre recombinase function in Xenopus embryos.
  JOURNAL   Nucleic Acids Res. 2001 Jun 1. 29(11):E53-3.
  PUBMED    11376165
REFERENCE   2  (bases 1 to 5416)
  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: "Nucleic 
            Acids Res. 2001 Jun 1. 29(11):E53-3."
COMMENT     Sequence Label: LCMV:ECFP(loxP)(FRT)MCS
FEATURES             Location/Qualifiers
     source          1..5416
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(34..67)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    75..108
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     regulatory      118..127
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(124..126,127..129,130..843)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(169..190)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(430..449)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     777..798
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    957..1091
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(1099..1132)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    1140..1173
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     intron          1298..1363
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     polyA_signal    1410..1544
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1698..1718)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1698..1716)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1737..1753)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1737..1753)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1750..1772)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1761..1777
                     /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(1785..1791,1792..1809,1810..1815))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1830..1851
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1968..1985)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2139..2727)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2219..2238)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2898..3689,3690..3758))
                     /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     3521..3540
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3759..3863)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3889..4344)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4026..4047)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4238..4257
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4432..5416
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttataactt cgtatagcat acattatacg
       61 aagttataac tagtgaagtt cctattctct agaaagtata ggaacttcag atccatcgcc
      121 accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
      181 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
      241 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
      301 accctcgtga ccaccctgac ctggggcgtg cagtgcttca gccgctaccc cgaccacatg
      361 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
      421 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
      481 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
      541 cacaagctgg agtacaacta catcagccac aacgtctata tcaccgccga caagcagaag
      601 aacggcatca aggccaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc
      661 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
      721 cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg
      781 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
      841 taatctagag gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa
      901 acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact
      961 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     1021 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     1081 atgtctggat ctgaattcat aacttcgtat agcatacatt atacgaagtt ataggatccg
     1141 aagttcctat tctctagaaa gtataggaac ttcccgggcc taggagatct caccggtgac
     1201 tagtcagggg ggatctgcat ggatctttgt gaaggaacct tacttctgtg gtgtgacata
     1261 attggacaaa ctacctacag agatttaaag ctctaaggta aatataaaat ttttaagtgt
     1321 ataatgtgtt aaactactga ttctaattgt ttgtgtattt tagattccaa cctatggaac
     1381 tgatgaatgg gagcagtggt ggaatgccag atccagacat gataagatac attgatgagt
     1441 ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg
     1501 ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca
     1561 ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc
     1621 tctacaaatg tggtatggct gattatgatc tgcggccgcg gcgccaatgc attgggcccg
     1681 gtacccagct tttgttccct ttagtgaggg ttaattgcgc gcttggcgta atcatggtca
     1741 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     1801 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     1861 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     1921 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     1981 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     2041 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     2101 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     2161 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     2221 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     2281 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     2341 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     2401 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     2461 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     2521 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg
     2581 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     2641 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     2701 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     2761 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     2821 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     2881 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     2941 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     3001 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     3061 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     3121 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     3181 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     3241 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     3301 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     3361 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     3421 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     3481 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     3541 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     3601 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     3661 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     3721 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     3781 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     3841 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctaa attgtaagcg
     3901 ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat
     3961 aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg
     4021 ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc
     4081 gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt
     4141 tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag
     4201 cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg
     4261 gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc
     4321 ttaatgcgcc gctacagggc gcgtcccatt cgccattcag gctgcgcaac tgttgggaag
     4381 ggcgatcggt gcgggcctct tcgctattac gccagtcgag gcggccgcgt cgaccatagc
     4441 caattcaata tggcgtatat ggactcatgc caattcaata tggtggatct ggacctgtgc
     4501 caattcaata tggcgtatat ggactcgtgc caattcaata tggtggatct ggaccccagc
     4561 caattcaata tggcggactt ggcaccatgc caattcaata tggcggactt ggcactgtgc
     4621 caactgggga ggggtctact tggcacggtg ccaagtttga ggaggggtct tggccctgtg
     4681 ccaagtccgc catattgaat tggcatggtg ccaataatgg cggccatatt ggctatatgc
     4741 caggatcaat atataggcaa tatccaatat ggccctatgc caatatggct attggccagg
     4801 ttcaatacta tgtattggcc ctatgccata tagtattcca tatatgggtt ttcctattga
     4861 cgtagatagc ccctcccaat gggcggtccc atataccata tatggggctt cctaataccg
     4921 cccatagcca ctcccccatt gacgtcaatg gtctctatat atggtctttc ctattgacgt
     4981 catatgggcg gtcctattga cgtatatggc gcctccccca ttgacgtcaa ttacggtaaa
     5041 tggcccgcct ggctcaatgc ccattgacgt caataggacc acccaccatt gacgtcaatg
     5101 ggatggctca ttgcccattc atatccgttc tcacgccccc tattgacgtc aatgacggta
     5161 aatggcccac ttggcagtac atcaatatct attaatagta acttggcaag tacattacta
     5221 ttggaagtac gccagggtac attggcagta ctcccattga cgtcaatggc ggtaaatggc
     5281 ccgcgatggc tgccaagtac atccccattg acgtcaatgg ggaggggcaa tgacgcaaat
     5341 gggcgttcca ttgacgtaaa tgggcggtag gcgtgcctaa tgggaggtct atataagcaa
     5401 tgctcgttta gggaac
//
