LOCUS       sequence-135638-        8117 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  promoterless Tol2 vector with 6xmyc tag upstream of and in frame 
            with attR1/attR2 cassette and late SV40 polyadenylation signal.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8117)
  AUTHORS   Villefranc JA, Amigo J, Lawson ND
  TITLE     Gateway compatible vectors for analysis of gene function in the 
            zebrafish.
  JOURNAL   Dev Dyn. 2007 Nov . 236(11):3077-87.
  PUBMED    17948311
REFERENCE   2  (bases 1 to 8117)
  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: "Dev Dyn. 
            2007 Nov . 236(11):3077-87."
COMMENT     Sequence Label: 252 pTol2-MTDest
FEATURES             Location/Qualifiers
     source          1..8117
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(34..462)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(144..165)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     356..375
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     589..611
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     603..620
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     604..620
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        630..648
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(778..789,790..952)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     CDS             1348..1377
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1387..1416
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1426..1455
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1465..1494
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1504..1533
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1567..1596
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     protein_bind    1624..1748
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1773..1778,1779..1796,1797..1803)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1857..2516
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1923..1942)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2858..3163
                     /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
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3042..3061
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(3204..3328)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     polyA_signal    3372..3506
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3396..3415)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3450..3469
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    join(5647..5834,5835..5846)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(5880..5896)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5880..5896)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(5933..5953)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5933..5951)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5972..5988)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5972..5988)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5985..6007)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5996..6012
                     /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(6020..6026,6027..6044,6045..6050))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6065..6086
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6203..6220)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6374..6962)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6454..6473)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7133..7924,7925..7993))
                     /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     7756..7775
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7994..8098)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
       61 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
      121 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
      181 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
      241 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
      301 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
      361 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
      421 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg ccattcaggc
      481 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
      541 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
      601 gttgtaaaac gacggccagt gagcgcgcgt aatacgactc actatagggc gaattgggta
      661 caggtcattg cacaacacca gaaatgccct ctgatctgca aaagacgtga atatctgttc
      721 agacacccat atccactctg ttccacacag gtcagaggtt tgtccaggag ttcttgacag
      781 aggtgtaaaa agtactcaaa aattttactc aagtgaaagt acaagtactt agggaaaatt
      841 ttactcaatt aaaagtaaaa gtatctggct agaatcttac ttgagtaaaa gtaaaaaagt
      901 actccattaa aattgtactt gagtattaag gaagtaaaag taaaagcaag aaagaaaact
      961 agagattctt gtttaagctt ttaatctcaa aaaacattaa atgaaatgca tacaaggttt
     1021 tatcctgctt tagaactgtt tgtatttaat tatcaaacta taagacagac aatctaatgc
     1081 cagtacacgc tactcaaagt tgtaaaacct cagatttaac ttcagtagaa gctgattctc
     1141 aaaattgtta gtgtcaagcc tagctctttt ggggctgaaa agcaatcctg cagtgctgaa
     1201 aagcctctca caggcagccg atgcgggaag aggtgtatta gtcttgatag agaggctgca
     1261 aatagcagga aacgtgagca gagactccct ggtgtctgaa acacaggcca gatgggccga
     1321 tatcgctagc atcgatttaa agctatggag caaaagctca tttctgaaga ggacttgaat
     1381 gaaatggagc aaaagctcat ttctgaagag gacttgaatg aaatggagca aaagctcatt
     1441 tctgaagagg acttgaatga aatggagcaa aagctcattt ctgaagagga cttgaatgaa
     1501 atggagcaaa agctcatttc tgaagaggac ttgaatgaaa tggagagctt gggcgacctc
     1561 accatggagc aaaagctcat ttctgaagag gacttgaatt tagagatctc gaattcaagg
     1621 atcacaagtt tgtacaaaaa agctgaacga gaaacgtaaa atgatataaa tatcaatata
     1681 ttaaattaga ttttgcataa aaaacagact acataatact gtaaaacaca acatatccag
     1741 tcatattggc ggccgcatta ggcaccccag gctttacact ttatgcttcc ggctcgtata
     1801 atgtgtggat tttgagttag gatccgtcga gattttcagg agctaaggaa gctaaaatgg
     1861 agaaaaaaat cactggatat accaccgttg atatatccca atggcatcgt aaagaacatt
     1921 ttgaggcatt tcagtcagtt gctcaatgta cctataacca gaccgttcag ctggatatta
     1981 cggccttttt aaagaccgta aagaaaaata agcacaagtt ttatccggcc tttattcaca
     2041 ttcttgcccg cctgatgaat gctcatccgg aattccgtat ggcaatgaaa gacggtgagc
     2101 tggtgatatg ggatagtgtt cacccttgtt acaccgtttt ccatgagcaa actgaaacgt
     2161 tttcatcgct ctggagtgaa taccacgacg atttccggca gtttctacac atatattcgc
     2221 aagatgtggc gtgttacggt gaaaacctgg cctatttccc taaagggttt attgagaata
     2281 tgtttttcgt ctcagccaat ccctgggtga gtttcaccag ttttgattta aacgtggcca
     2341 atatggacaa cttcttcgcc cccgttttca ccatgggcaa atattatacg caaggcgaca
     2401 aggtgctgat gccgctggcg attcaggttc atcatgccgt ttgtgatggc ttccatgtcg
     2461 gcagaatgct taatgaatta caacagtact gcgatgagtg gcagggcggg gcgtaaacgc
     2521 gtggatccgg cttactaaaa gccagataac agtatgcgta tttgcgcgct gatttttgcg
     2581 gtataagaat atatactgat atgtataccc gaagtatgtc aaaaagaggt atgctatgaa
     2641 gcagcgtatt acagtgacag ttgacagcga cagctatcag ttgctcaagg catatatgat
     2701 gtcaatatct ccggtctggt aagcacaacc atgcagaatg aagcccgtcg tctgcgtgcc
     2761 gaacgctgga aagcggaaaa tcaggaaggg atggctgagg tcgcccggtt tattgaaatg
     2821 aacggctctt ttgctgacga gaacaggggc tggtgaaatg cagtttaagg tttacaccta
     2881 taaaagagag agccgttatc gtctgtttgt ggatgtacag agtgatatta ttgacacgcc
     2941 cgggcgacgg atggtgatcc ccctggccag tgcacgtctg ctgtcagata aagtctcccg
     3001 tgaactttac ccggtggtgc atatcgggga tgaaagctgg cgcatgatga ccaccgatat
     3061 ggccagtgtg ccggtctccg ttatcgggga agaagtggct gatctcagcc accgcgaaaa
     3121 tgacatcaaa aacgccatta acctgatgtt ctggggaata taaatgtcag gctcccttat
     3181 acacagccag tctgcaggtc gaccatagtg actggatatg ttgtgtttta cagcattatg
     3241 tagtctgttt tttatgcaaa atctaattta atatattgat atttatatca ttttacgttt
     3301 ctcgttcagc tttcttgtac aaagtggtga tcctctcgag cctctagaac tatagtgagt
     3361 cgtattacgt agatccagac atgataagat acattgatga gtttggacaa accacaacta
     3421 gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa
     3481 ccattataag ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg
     3541 ttcaggggga ggtgtgggag gttttttaat tcgcggccgc ggcgccaatg cattgggccc
     3601 tcgagcatat gagatctgtc ccgatctttc tcttctgtgc tgtcaattta gagtagtttt
     3661 tgaggtaatt tgggtgcatt ctctgtaaaa acaacaaata aatatttata ttgtatgcaa
     3721 taaatgtaac aattacacat acactctctg attacagcat taaagggtta gttcacccaa
     3781 aaatgaaaag atgtcattaa tgactcaccc tcatgtcgtt ccaagcccgt aacacctccg
     3841 ttcatcttcg gaacacagtt taagatattt tagatttagt ccgagagctt tctgtccctc
     3901 cactgaaaat gtatgtacgg tatactgtcc atgtccagaa aggtaataaa aacatcaaag
     3961 tagtccatgt gacatcagtg ggttaggtag aattatttga agcatcgaaa atacattttg
     4021 gtccaaaaat aacaaaacct acgactttat tcagcattgt agcgtcactg cgaagtcgtg
     4081 aacgcggatt gacaacagac ccggaagaga atacaatgcc gaataaagtc gtaggttttg
     4141 ttatttttgg accaaaatgt attcgatgct tcaaaaaatt ctaactaacc cactgatgtc
     4201 acatggacta ctttgatgtt tttattacct ttctggacat ggacagtata ccgtacatac
     4261 attctcaatg gaggcacaga aagctctcgg actaaatcta aaatatctta aactgtgttc
     4321 tgaagatgaa cggaggtctt acgggcttgg aacgacatga gggcgagtca ttaatgacat
     4381 tattttcatt tttgggtgaa ctaacccttt aataaaactg acatttgcag ccaattgcac
     4441 actttataat gttaaatact tgttgatttt agaaatacag tagcatttag tgacaatcaa
     4501 acattaattt tatttctgat tatattgaca aaaacacagg actgtaaggc tgacatgaca
     4561 gggtgaattg atttcattac attagttttg atggtaacac ttttacaata aggttcgtta
     4621 gttaactaca ttagttaaca tgaactaata atgaactgca cgtatacatg ttaatttcaa
     4681 catttactaa tactttatat cttgttaaca ttagttaatg cactgtgaac taacatgaac
     4741 aaacaatgaa catctgtatt tctattaact aacgttaaca aagattaaca gatacagtaa
     4801 caaatgtatt gctcatggtt atttaatgtt agttaataca tgaactaatc aacctcattg
     4861 taaagtgtta ccgttttgac aatacaacat gtttggaaaa aaatttctcg cttggttttc
     4921 ttgtgttttg catgaattca aacacaatga gagcacggaa tgataatgat tattcaaagc
     4981 tgcaataggc gccaaatttc ccgtggacct aacgttacca attacaaatg cattttagca
     5041 ttttattacg tgagccttat attgtcaatc gcatgcatta gtgaaaatag gcctactaca
     5101 catataaaag gcgcgccatc aacgtcaaaa catgcacacc caaaaaaaaa aaaaaaaaaa
     5161 aaaaaaaaaa ctatcagtaa aacaaaatac ttaatgtact tacctcaata tgcttcctta
     5221 ggtttgatgg cgaacttttg aaggccgata tttctttatt aagcgggaga cagaggacac
     5281 atttcatctt gaatgaatct ttatttacac cacttaaaga aaagaattcg cgaagatacg
     5341 gccacgggtg ctcttgatcc tgtggctgat tttggactgt gctgctcgca gctgctgatg
     5401 aatcacatac ttcctccatt ttcttccact gattgactgt tataatttcc ctaatttcca
     5461 ggtcaaggtg ctgtgcattg tggtaataga tgtgacatga cgtcacttcc aaaggaccaa
     5521 tgaacatgtc tgaccaattt catataatgt gaaaacgatt ttcataggca gaataaataa
     5581 catttaaatt aaactgggca tcagcgcaat tcaattggtt tggtaatagc aagggaaaat
     5641 agaatgaagt gatctccaaa aaataagtac tttttgactg taaataaaat tgtaaggagt
     5701 aaaaagtact tttttttcta aaaaaatgta attaagtaaa agtaaaagta ttgattttta
     5761 attgtactca agtaaagtaa aaatccccaa aaataatact taagtacagt aatcaagtaa
     5821 aattactcaa gtactttaca cctctggttc ttgaccccct accttcagca agcccagcag
     5881 atccactagt tctagagcgg ccgccaccgc ggtggagctc cagcttttgt tccctttagt
     5941 gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     6001 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     6061 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     6121 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     6181 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     6241 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     6301 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     6361 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     6421 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     6481 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     6541 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     6601 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     6661 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     6721 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     6781 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     6841 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     6901 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     6961 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     7021 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     7081 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     7141 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     7201 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     7261 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     7321 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     7381 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     7441 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     7501 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     7561 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     7621 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     7681 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     7741 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     7801 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     7861 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     7921 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     7981 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     8041 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     8101 catttccccg aaaagtg
//
