LOCUS       sequence-14295-e        5828 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 5828)
  AUTHORS   Waldner C, Rempel O, Schutte F, Yanik M, Solomentsew N, Ryffel GU
  TITLE     Double conditional human embryonic kidney cell line based on FLP and
            PhiC31 mediated transgene integration.
  JOURNAL   BMC Res Notes. 2011 Oct 18;4:420.
  PUBMED    22008483
REFERENCE   2  (bases 1 to 5828)
  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: "BMC Res 
            Notes."; date: "2011-10-18"; volume: "4"; pages: "420"
COMMENT     Sequence Label: pDOCKING-Neo
FEATURES             Location/Qualifiers
     source          1..5828
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             join(52..54,55..765)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(94..115)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(355..374)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     702..723
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             772..1566
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(826..845)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1436..1455
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1740..1861
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1777..1796)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1831..1850
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1906..1926)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1906..1924)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1945..1961)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1945..1961)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1958..1980)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1969..1985
                     /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(1993..1999,2000..2017,2018..2023))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2038..2059
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2176..2193)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2347..2935)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2427..2446)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3106..3897,3898..3966))
                     /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     3729..3748
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3967..4071)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4097..4552)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4234..4255)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4446..4465
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4679..4701
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4693..4710
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4694..4710
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4720..4739
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4720..4738
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        4771..5755
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     protein_bind    complement(5789..5822)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
ORIGIN
        1 atgtccccaa ctggggtaac ctttgagttc tctcagttgg gggcgagatc tgtgagcaag
       61 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
      121 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
      181 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
      241 ctgacctggg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
      301 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
      361 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
      421 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
      481 aactacatca gccacaacgt ctatatcacc gccgacaagc agaagaacgg catcaaggcc
      541 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
      601 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
      661 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
      721 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaaggaatt catgattgaa
      781 caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac
      841 tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg
      901 cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag
      961 gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt
     1021 gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg
     1081 tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg
     1141 catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga
     1201 gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag
     1261 gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat
     1321 ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt
     1381 tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg
     1441 gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt
     1501 tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc
     1561 ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac
     1621 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
     1681 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca
     1741 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     1801 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     1861 atcatgtctg cggccgccac cgcggtggag ctccagcttt tgttcccttt agtgagggtt
     1921 aattgcgcgc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     1981 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     2041 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     2101 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     2161 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     2221 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     2281 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     2341 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     2401 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     2461 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     2521 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     2581 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     2641 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     2701 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     2761 gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc
     2821 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     2881 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     2941 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     3001 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     3061 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     3121 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     3181 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     3241 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     3301 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     3361 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     3421 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     3481 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     3541 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     3601 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     3661 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     3721 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     3781 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     3841 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     3901 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     3961 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     4021 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     4081 ccgaaaagtg ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt
     4141 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca
     4201 aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta
     4261 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
     4321 cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
     4381 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
     4441 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg
     4501 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg
     4561 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc
     4621 cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc
     4681 cagtcacgac gttgtaaaac gacggccagt gagcgcgcgt aatacgactc actatagggc
     4741 gaattgggta ccgggccccc cctcgaggtc gaccatagcc aattcaatat ggcgtatatg
     4801 gactcatgcc aattcaatat ggtggatctg gacctgtgcc aattcaatat ggcgtatatg
     4861 gactcgtgcc aattcaatat ggtggatctg gaccccagcc aattcaatat ggcggacttg
     4921 gcaccatgcc aattcaatat ggcggacttg gcactgtgcc aactggggag gggtctactt
     4981 ggcacggtgc caagtttgag gaggggtctt ggccctgtgc caagtccgcc atattgaatt
     5041 ggcatggtgc caataatggc ggccatattg gctatatgcc aggatcaata tataggcaat
     5101 atccaatatg gccctatgcc aatatggcta ttggccaggt tcaatactat gtattggccc
     5161 tatgccatat agtattccat atatgggttt tcctattgac gtagatagcc cctcccaatg
     5221 ggcggtccca tataccatat atggggcttc ctaataccgc ccatagccac tcccccattg
     5281 acgtcaatgg tctctatata tggtctttcc tattgacgtc atatgggcgg tcctattgac
     5341 gtatatggcg cctcccccat tgacgtcaat tacggtaaat ggcccgcctg gctcaatgcc
     5401 cattgacgtc aataggacca cccaccattg acgtcaatgg gatggctcat tgcccattca
     5461 tatccgttct cacgccccct attgacgtca atgacggtaa atggcccact tggcagtaca
     5521 tcaatatcta ttaatagtaa cttggcaagt acattactat tggaagtacg ccagggtaca
     5581 ttggcagtac tcccattgac gtcaatggcg gtaaatggcc cgcgatggct gccaagtaca
     5641 tccccattga cgtcaatggg gaggggcaat gacgcaaatg ggcgttccat tgacgtaaat
     5701 gggcggtagg cgtgcctaat gggaggtcta tataagcaat gctcgtttag ggaaccgcca
     5761 ttctgcctgg ggacgtcgga gcaagcttat aacttcgtat agcatacatt atacgaagtt
     5821 ataggatc
//
