LOCUS       sequence-11441-e        4032 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 4032)
  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 4032)
  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-TAP
FEATURES             Location/Qualifiers
     source          1..4032
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      517..526
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(529..562)
                     /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)."
     CDS             857..874
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             902..922
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             929..994
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    1184..1318
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1221..1240)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1275..1294
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1336..1383
                     /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(1407..1423)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1407..1423)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1420..1442)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1431..1447
                     /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(1455..1461,1462..1479,1480..1485))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1500..1521
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1638..1655)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1809..2397)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1889..1908)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2568..3359,3360..3428))
                     /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     3191..3210
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3429..3533)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3601..3619
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3657..3679)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3779..3798
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3992..4014
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4006..4023
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4007..4023
                     /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 gtcggcgaag
      361 ccggtgagtg agcggcgcgg ggccaatcgc gtgcgccgtt ccgaaagttg ccttttatgg
      421 ctcgagcggc cgcggcggcg ccctataaaa cccagcggcg cgacgcgcca ccaccgccga
      481 gaccgcgtcc gcccgcgagc acagagcctc gcctttgcca ccatggctat aacttcgtat
      541 agcatacatt atacgaacgg tagtgatagg ccttttagct acatctgcca atccatctca
      601 ttttcacaca cacacacacc actttccttc tggtcagtgg gcacatgtcc agcctcaagt
      661 ttatatcacc acccccaatg cccaacactt gtatggcctt gggcgggtca tccccccccc
      721 cacccccagt atctgcaacc tcaagctagc ttgggtgcgt tggttgtgga taagtagcta
      781 gactccagca accagtaacc tctgcccttt ctcctccatg acaaccaggt accggtgcat
      841 gcggcgcgcc actagtcatc accatcacca tcacgattat gatattccaa ctactgctag
      901 cgagaatttg tattttcagg gtgagctcga ctacaaagac catgacggtg attataaaga
      961 tcatgacatc gactacaagg atgacgatga caagtaggat atcgctagcg gatccaccac
     1021 ctacagccca gtggggtcgc tctagagcga cctcgagggg gggctagata agtaaatgat
     1081 cataatcagc catatcacat ctgtagaggt tttacttgct ttaaaaaacc tcccacacct
     1141 ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc
     1201 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     1261 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctagatccg
     1321 ggggtaccgc gtcgagaagt tcctattccg aagttcctat tctctagaaa gtataggaac
     1381 ttcgtcgacg cgaattcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     1441 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     1501 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     1561 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     1621 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     1681 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     1741 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     1801 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     1861 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     1921 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     1981 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     2041 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     2101 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     2161 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     2221 tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag
     2281 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     2341 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     2401 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     2461 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     2521 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     2581 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     2641 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     2701 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     2761 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     2821 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     2881 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     2941 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     3001 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     3061 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     3121 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     3181 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     3241 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     3301 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     3361 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     3421 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     3481 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     3541 ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa
     3601 aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc
     3661 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
     3721 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct taactatgcg
     3781 gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc gcacagatgc
     3841 gtaaggagaa aataccgcat caggcgccat tcgccattca ggctgcgcaa ctgttgggaa
     3901 gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca
     3961 aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc
     4021 agtgccaagc tt
//
