LOCUS       sequence-9309-ex        5880 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 5880)
  AUTHORS   Voutev R, Hubbard EJ
  TITLE     A ""FLP-Out"" system for controlled gene expression in 
            Caenorhabditis elegans.
  JOURNAL   Genetics. 2008 Sep;180(1):103-19. doi: 10.1534/genetics.108.090274.
  PUBMED    18723890
REFERENCE   2  (bases 1 to 5880)
  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"; doi: 
            "10.1534/genetics.108"; journalName: "Genetics"; date: "2008-09"; 
            volume: "180"; issue: "1"; pages: "103-19"
COMMENT     Sequence Label: pGC319
FEATURES             Location/Qualifiers
     source          1..5880
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    91..214
                     /gene="mutant version of attR"
                     /label=attR4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(255..560)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(357..376)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(902..1561)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1476..1495
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(1615..1621,1622..1639,1640..1645))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(1671..1794)
                     /gene="mutant version of attR"
                     /label=attR3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(1819..1866)
                     /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)."
     CDS             1942..1962
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1984..2004
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2026..2046
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2068..2088
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2101..2811
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2246..2264)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2514..2533
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(3320..3342)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3442..3461
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3539..3557)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3625..3729
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3730..3798,3799..4590)
                     /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     complement(3948..3967)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4761..5349
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5250..5269
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5503..5520
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5637..5658
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5673..5678,5679..5696,5697..5703)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5711..5727
                     /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)."
     primer_bind     5716..5738
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5735..5751
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5735..5751
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 aagcttgcat gcctgcaggt cgactctaga ccgcggtttt ggcgcggatc cactagttct
       61 agcttcaaaa gcctcattac gccaagctat caactttgta tagaaaagtt gaacgagaaa
      121 cgtaaaatga tataaatatc aatatattaa attagatttt gcataaaaaa cagactacat
      181 aatactgtaa aacacaacat atccagtcac tatggtcgac ctgcagactg gctgtgtata
      241 agggagcctg acatttatat tccccagaac atcaggttaa tggcgttttt gatgtcattt
      301 tcgcggtggc tgagatcagc cacttcttcc ccgataacgg agaccggcac actggccata
      361 tcggtggtca tcatgcgcca gctttcatcc ccgatatgca ccaccgggta aagttcacgg
      421 gggactttat ctgacagcag acgtgcactg gccaggggga tcaccatccg tcgcccgggc
      481 gtgtcaataa tatcactctg tacatccaca aacagacgat aacggctctc tcttttatag
      541 gtgtaaacct taaactgcat ttcaccagcc cctgttctcg tcggcaaaag agccgttcat
      601 ttcaataaac cgggcgacct cagccatccc ttcctgattt tccgctttcc agcgttcggc
      661 acgcagacga cgggcttcat tctgcatggt tgtgcttacc gaaccggaga tattgacatc
      721 atatatgcct tgagcaactg atagctgtcg ctgtcaactg tcactgtaat acgctgcttc
      781 atagcatacc tctttttgac atacttcggg tatacatatc agtatatatt cttataccgc
      841 aaaaatcagc gcgcaaatac gcatactgtt atctggcttt tagtaagccg gatcctctag
      901 attacgcccc gccctgccac tcatcgcagt actgttgtaa ttcattaagc attctgccga
      961 catggaagcc atcacaaacg gcatgatgaa cctgaatcgc cagcggcatc agcaccttgt
     1021 cgccttgcgt ataatatttg cccatggtga aaacgggggc gaagaagttg tccatattgg
     1081 ccacgtttaa atcaaaactg gtgaaactca cccagggatt ggctgagacg aaaaacatat
     1141 tctcaataaa ccctttaggg aaataggcca ggttttcacc gtaacacgcc acatcttgcg
     1201 aatatatgtg tagaaactgc cggaaatcgt cgtggtattc actccagagc gatgaaaacg
     1261 tttcagtttg ctcatggaaa acggtgtaac aagggtgaac actatcccat atcaccagct
     1321 caccgtcttt cattgccata cggaattccg gatgagcatt catcaggcgg gcaagaatgt
     1381 gaataaaggc cggataaaac ttgtgcttat ttttctttac ggtctttaaa aaggccgtaa
     1441 tatccagctg aacggtctgg ttataggtac attgagcaac tgactgaaat gcctcaaaat
     1501 gttctttacg atgccattgg gatatatcaa cggtggtata tccagtgatt tttttctcca
     1561 ttttagcttc cttagctcct gaaaatctcg acggatccta actcaaaatc cacacattat
     1621 acgagccgga agcataaagt gtaaagcctg ggggtgccta atgcggccgc catagtgact
     1681 ggatatgttg tgttttacag tattatgtag tctgtttttt atgcaaaatc taatttaata
     1741 tattgatatt tatatcattt tacgtttctc gttcaacttt attatacata gttgataatt
     1801 cactgggcta gccgctctga agttcctata ctttctagag aataggaact tcggaatagg
     1861 aacttcaaga tccccgggta tgtttcgaat gatactaaca taacatagaa cattttcagg
     1921 agctcagaaa aaatgactgc tccaaagaag aagcgtaagg taccggcaga aaaaatgact
     1981 gctccaaaga agaagcgtaa ggtaccggca gaaaaaatga ctgctccaaa gaagaagcgt
     2041 aaggtaccgg cagaaaaaat gactgctcca aagaagaagc gtaaggtacc ggtagaaaaa
     2101 atggtgagca agggcgagga ggataacatg gccatcatca aggagttcat gcgcttcaag
     2161 gtgcacatgg agggctccgt gaacggccac gagttcgaga tcgagggcga gggcgagggc
     2221 cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc
     2281 ttcgcctggg acatcctgtc ccctcagttc atgtacggct ccaaggccta cgtgaagcac
     2341 cccgccgaca tccccgacta cttgaagctg tccttccccg agggcttcaa gtgggagcgc
     2401 gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc aggactcctc cctgcaggac
     2461 ggcgagttca tctacaaggt gaagctgcgc ggcaccaact tcccctccga cggccccgta
     2521 atgcagaaga agaccatggg ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc
     2581 gccctgaagg gcgagatcaa gcagaggctg aagctgaagg acggcggcca ctacgacgct
     2641 gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc
     2701 aacatcaagt tggacatcac ctcccacaac gaggactaca ccatcgtgga acagtacgaa
     2761 cgcgccgagg gccgccactc caccggcggc atggacgagc tgtacaagta agaattcgct
     2821 agccggccat acaagtaatc cggatgatcg acgccaacgt cgttgaattt tcaaatttta
     2881 aatactgaat atttgttttt tttcctatta tttatttatt ctctttgtgt tttttttctt
     2941 gctttctaaa aaattaattc aatccaaatc taaacatgag cggttttttt tctctttccg
     3001 tctcccaatt cgtattccgc tcctctcatc tgaacacaat gtgcaagttt atttatcttc
     3061 tcgctttcat ttcattagga cgtgggggga attggtggaa gggggaaaca cacaaaagga
     3121 tgatggaaat gaaataagga cacacaatat gcaacaacat tcaattcaga aatatggagg
     3181 aaggtttaaa agaaaacata aaaatatata gaggaggaag gaaaactagt aaaaaataag
     3241 caaagaaatt aggcgaacga tgagaattgt cctcgcttgg gccctttcgt ctcgcgcgtt
     3301 tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc
     3361 tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt
     3421 gtcggggctg gcttaactat gcggcatcag agcagattgt actgagagtg caccatatgc
     3481 ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgg ccttaagggc
     3541 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     3601 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     3661 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     3721 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     3781 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     3841 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     3901 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     3961 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     4021 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     4081 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     4141 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     4201 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     4261 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     4321 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     4381 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     4441 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     4501 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     4561 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     4621 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     4681 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     4741 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     4801 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     4861 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     4921 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     4981 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     5041 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     5101 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gcattgagaa
     5161 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     5221 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     5281 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     5341 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     5401 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     5461 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     5521 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
     5581 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
     5641 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
     5701 ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac
     5761 gccaagctgt aagtttaaac atgatcttac taactaacta ttctcattta aattttcaga
     5821 gcttaaaaat ggctgaaatc actcacaacg atggatacgc taacaacttg gaaatgaaat
//
