LOCUS       sequence-11470-e        4512 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 4512)
  AUTHORS   Singla V, Hunkapiller J, Santos N, Seol AD, Norman AR, Wakenight P, 
            Skarnes WC, Reiter JF
  TITLE     Floxin, a resource for genetically engineering mouse ESCs.
  JOURNAL   Nat Methods. 2010 Jan . 7(1):50-2.
  PUBMED    19966808
REFERENCE   2  (bases 1 to 4512)
  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 
            Methods. 2010 Jan . 7(1):50-2."
COMMENT     Sequence Label: pFloxin-MCS2-IRES-MCS
FEATURES             Location/Qualifiers
     source          1..4512
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      520..529
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(532..565)
                     /label=lox66
                     /bound_moiety="Cre recombinase"
                     /note="Right element (RE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    879..1451
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    902..1453
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1046..1063)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     polyA_signal    1664..1798
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1701..1720)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1755..1774
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1816..1863
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     complement(1887..1903)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1887..1903)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1900..1922)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1911..1927
                     /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(1935..1941,1942..1959,1960..1965))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1980..2001
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2118..2135)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2289..2877)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2369..2388)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3048..3839,3840..3908))
                     /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     3671..3690
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3909..4013)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4081..4099
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4137..4159)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4259..4278
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4472..4494
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4486..4503
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4487..4503
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 aagcttcctt ctgcaggagc gtacagaacc cagggccctg gcacccgtgc agaccctggc
       61 ccaccccacc tgggcgctca gtgcccaaga gatgtccaca cctaggatgt cccgcggtgg
      121 gtggggggcc cgagagacgg gcaggccggg ggcaggcctg gccatgcggg gccgaaccgg
      181 gcactgccca gcgtggggcg cgggggccac ggcgcgcgcc cccagccccc gggcccagca
      241 ccccaaggcg gccaacgcca aaactctccc tcctcctctt cctcaatctc gctctcgctc
      301 tttttttttt tcgcaaaagg aggggagagg gggtaaaaaa atgctgcact gtgcggcgaa
      361 gccggtgagt gagcggcgcg gggccaatca gcgtgcgccg ttccgaaagt tgccttttat
      421 ggctcgagcg gccgcggcgg cgccctataa aacccagcgg cgcgacgcgc caccaccgcc
      481 gagaccgcgt ccgccccgcg agcacagagc ctcgcctttg ccaccatggc tataacttcg
      541 tatagcatac attatacgaa cggtagtgat aggcctttta gctacatctg ccaatccatc
      601 tcattttcac acacacacac accactttcc ttctggtcag tgggcacatg tccagcctca
      661 agtttatatc accaccccca atgcccaaca cttgtatggc cttgggcggg tcatcccccc
      721 ccccaccccc agtatctgca acctcaagct agcttgggtg cgttggttgt ggataagtag
      781 ctagactcca gcaaccagta acctctgccc tttctcctcc atgacaacca ggtaccggta
      841 gtgactaata gctgatagaa ttccgccccc cccccccccc cctctccctc ccccccccct
      901 aacgttactg gccgaagccg cttggaataa ggccggtgtg cgtttgtcta tatgttattt
      961 tccaccatat tgccgtcttt tggcaatgtg agggcccgga aacctggccc tgtcttcttg
     1021 acgagcattc ctaggggtct ttcccctctc gccaaaggaa tgcaaggtct gttgaatgtc
     1081 gtgaaggaag cagttcctct ggaagcttct tgaagacaaa caacgtctgt agcgaccctt
     1141 tgcaggcagc ggaacccccc acctggcgac aggtgcctct gcggccaaaa gccacgtgta
     1201 taagatacac ctgcaaaggc ggcacaaccc cagtgccacg ttgtgagttg gatagttgtg
     1261 gaaagagtca aatggctctc ctaagcgtat tcaacaaggg gctgaaggat gcccagaagg
     1321 taccccattg tatgggatct gatctggggc ctcggtgcac atgctttaca tgtgtttagt
     1381 cgaggttaaa aaaacgtcta ggccccccga accacgggga cgtggttttc ctttgaaaaa
     1441 cacgatgata ataggatcgc atgcggcgcg ccactagtga tatcgctagc ggatccacca
     1501 cctacagccc agtggggtcg ctctagagcg acctcgaggg ggggctagat aagtaatgat
     1561 cataatcagc catatcacat ctgtagaggt tttacttgct ttaaaaaacc tcccacacct
     1621 ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc
     1681 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     1741 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctagatccg
     1801 ggggtaccgc gtcgagaagt tcctattccg aagttcctat tctctagaaa gtataggaac
     1861 ttcgtcgacg cgaattcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     1921 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     1981 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     2041 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     2101 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     2161 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     2221 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     2281 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     2341 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     2401 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     2461 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     2521 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     2581 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     2641 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     2701 tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag
     2761 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     2821 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     2881 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     2941 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     3001 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     3061 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     3121 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     3181 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     3241 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     3301 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     3361 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     3421 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     3481 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     3541 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     3601 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     3661 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     3721 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     3781 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     3841 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     3901 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     3961 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     4021 ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa
     4081 aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc
     4141 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
     4201 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct taactatgcg
     4261 gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc gcacagatgc
     4321 gtaaggagaa aataccgcat caggcgccat tcgccattca ggctgcgcaa ctgttgggaa
     4381 gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca
     4441 aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc
     4501 agtgccaagc tt
//
