LOCUS       sequence-57047-e        7574 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 7574)
  AUTHORS   Tryon RC, Pisat N, Johnson SL, Dougherty JD
  TITLE     Development of translating ribosome affinity purification for 
            zebrafish.
  JOURNAL   Genesis. 2012 Dec 19. doi: 10.1002/dvg.22363.
  PUBMED    23281262
REFERENCE   2  (bases 1 to 7574)
  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: "Genesis. 
            2012 Dec 19. doi: 10.1002/dvg.22363."
COMMENT     Sequence Label: Tol2-zTRAP
FEATURES             Location/Qualifiers
     source          1..7574
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    10..144
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(34..53)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     88..107
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    join(583..770,771..782)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(816..832)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(816..832)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(869..889)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(869..887)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(908..924)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(908..924)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(921..943)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    932..948
                     /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(956..962,963..980,981..986))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1001..1022
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1139..1156)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1310..1898)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1390..1409)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2069..2860,2861..2929))
                     /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     2692..2711
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2930..3034)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3060..3515)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3197..3218)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3409..3428
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3642..3664
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3656..3673
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3657..3673
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3683..3702
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3683..3701
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(5587..5598,5599..5761)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     regulatory      6164..6173
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(6170..6172,6173..6175,6176..6886)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(6170..6172,6173..6175,6176..6886)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(6215..6236)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(6476..6495)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6823..6844
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     RBS             6961..6969
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
ORIGIN
        1 atcgatgatg atccagacat gataagatac attgatgagt ttggacaaac cacaactaga
       61 atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc
      121 attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt
      181 cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtatggct
      241 gattatgatc ctctagatca gatctgcgaa gatacggcca cgggtgctct tgatcctgtg
      301 gctgattttg gactgtgctg ctcgcagctg ctgatgaatc acatacttcc tccattttct
      361 tccactgatt gactgttata atttccctaa tttccaggtc aaggtgctgt gcattgtggt
      421 aatagatgtg acatgacgtc acttccaaag gaccaatgaa catgtctgac caatttcata
      481 taatgtgaaa acgattttca taggcagaat aaataacatt taaattaaac tgggcatcag
      541 cgcaattcaa ttggtttggt aatagcaagg gaaaatagaa tgaagtgatc tccaaaaaat
      601 aagtactttt tgactgtaaa taaaattgta aggagtaaaa agtacttttt tttctaaaaa
      661 aatgtaatta agtaaaagta aaagtattga tttttaattg tactcaagta aagtaaaaat
      721 ccccaaaaat aatacttaag tacagtaatc aagtaaaatt actcaagtac tttacacctc
      781 tggttcttga ccccctacct tcagcaagcc cagcagatcc actagttcta gagcggccgc
      841 caccgcggtg gagctccagc ttttgttccc tttagtgagg gttaattgcg cgcttggcgt
      901 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
      961 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     1021 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     1081 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     1141 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     1201 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     1261 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     1321 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     1381 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     1441 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     1501 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     1561 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     1621 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     1681 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     1741 acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     1801 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     1861 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     1921 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     1981 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     2041 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     2101 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     2161 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     2221 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     2281 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     2341 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     2401 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     2461 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     2521 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     2581 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     2641 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     2701 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     2761 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     2821 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     2881 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     2941 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     3001 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccaccta
     3061 aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt
     3121 ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat
     3181 agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa
     3241 cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta
     3301 atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc
     3361 ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc
     3421 gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac
     3481 acccgccgcg cttaatgcgc cgctacaggg cgcgtcccat tcgccattca ggctgcgcaa
     3541 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg
     3601 atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa
     3661 aacgacggcc agtgagcgcg cgtaatacga ctcactatag ggcgaattgg gtaccaaata
     3721 gtaggaatta cccacctgta caagtgctga aaacttggat gaataagccc gtttgccatt
     3781 ttctactgct attttaaatc ttttctgtat ttgtctgcat ttgtctttac ccttgaataa
     3841 atgttttagt tgtttttttt caattatgac tgtgtttaac agacattatt acaataatat
     3901 gtaatagtag tacacactat tattatgtaa tagtacttgt tgactgtatt tgagaactgg
     3961 accgagtgag tgttacgtca cccattcaaa atgacttact tctggctcca acaaaatgaa
     4021 gttaattcag ttgccatttt tcactgtatg gacatcgccg tgttggagct agacgtcgcc
     4081 aataagcaag aaagagccga gatgcgtcga gtctgagtca ccgttcctat ggcaacccct
     4141 ctaaccaatc agaagtaagc ttgttggaag tccacagcct accacttgaa agcgggctgc
     4201 acaaaatctg tcaaacgttt tgaacgttgg atgtgagagc acatactttt attaaggcat
     4261 ctggttggtc agtttataat atcaacaact tgggctacag aaaagaaaag ttattacaga
     4321 aattatgatt aacaagtaca tgttaaataa agattttaat atgaatgcca ccactggagc
     4381 attcatgcca tttggagctt cttcctgttt ggatcactag aaggaggagg tcactcatta
     4441 cagttctcat atacagtcgt tggttggttg gttggttggt tggttggtag attgattgat
     4501 tgattgattg attgattgat tgattgattg attgattgat tgattgattg attgattgat
     4561 tgattgattg attgattgat tgattgattg attggtagtc aaaataagaa ataatttcca
     4621 cagattcatt acagaaatga ttaaatgcat acataaaaaa ctgggggggg ggggatacaa
     4681 caacacactt aagtcacatt tgcctacgta aagaaaagta aagaaaatca atagctatat
     4741 tttacatctc ctttttttgc tgtcttttaa ttagccttgt tttgctgtta tcttgattga
     4801 aactgtaact tcttcacctg cttcttttct ttgtaggttt tgccagccca gaaacaggca
     4861 tggctgtgca aggccagagc accatgcaca atgcgctgca tgtcttcatg aacggctcaa
     4921 tgtcctcagt ccagggctca gccaacgacc ccatcttcct ccttcaccat gctttcattg
     4981 acaggtaaca aacacgtcat gacattagac tgcacagttt ttgacaaagt tcatacaatc
     5041 tgttgtttat agctgctaca attagtgaag tttgtgaatg tacttggatg agcagcgaaa
     5101 gatcaattga gatcaattgt tagagtttgg ttgccctgca gagcaaagaa caaaaaataa
     5161 tctggtggct ttactgcgtg aggttattat tggtggaata gaaacacaaa acataattgc
     5221 atttatttgt ttaatttttt atcttatctt aactttcatc ttgcatattt gtttcttaca
     5281 tcatttctag catctttgag cgctggctaa gaactcatca gcctccccgg tccatctacc
     5341 cacgtaccaa tgcaccaatt ggccacaatg acggctacta catggtgcca ttccttcctc
     5401 tttataggaa tggagactac ctcctgtcca acaaggctct tggatacgag tacgcctacc
     5461 tgttggaccc aggtcattgc acaacaccag aaatgccctc tgatctgcaa aagacgtgaa
     5521 tatctgttca gacacccata tccactctgt tccacacagg tcagaggttt gtccaggagt
     5581 tcttgacaga ggtgtaaaaa gtactcaaaa attttactca agtgaaagta caagtactta
     5641 gggaaaattt tactcaatta aaagtaaaag tatctggcta gaatcttact tgagtaaaag
     5701 taaaaaagta ctccattaaa attgtacttg agtattaagg aagtaaaagt aaaagcaaga
     5761 aagaaaacta gagattcttg tttaagcttt taatctcaaa aaacattaaa tgaaatgcat
     5821 acaaggtttt atcctgcttt agaactgttt gtatttaatt atcaaactat aagacagaca
     5881 atctaatgcc agtacacgct actcaaagtt gtaaaacctc agatttaact tcagtagaag
     5941 ctgattctca aaattgttag tgtcaagcct agctcttttg gggctgaaaa gcaatcctgc
     6001 agtgctgaaa agcctctcac aggcagccga tgcgggaaga ggtgtattag tcttgataga
     6061 gaggctgcaa atagcaggaa acgtgagcag agactccctg gtgtctgaaa cacaggccag
     6121 atgggccctc gagtgctagc atgtcgacag ggatccaccg gtcgccacca tggtgagcaa
     6181 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
     6241 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
     6301 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
     6361 cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt
     6421 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
     6481 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
     6541 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
     6601 caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt
     6661 gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca
     6721 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac
     6781 ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     6841 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtccg gccggactca
     6901 gatctcgagc tcaagcttcg aattcagcaa ggtctcgaga gacacgttgt acgaggccgt
     6961 taaggaggtc caggccggtt cccgtcgcaa gaagagaaag tttcttgaaa ccgtggaact
     7021 ccagatcagc ttgaagaact acgaccctca gaaggacaag cgtttctcag gcactgtcag
     7081 gctgaagacc accccgcggc ccaagttctc tgtctgcgtt cttggagacc agcagcattg
     7141 tgatgaagca aaggccgctg aactgccaca catggacatt gaggctctta agaagctcaa
     7201 taagaacaag aaactggtca agaagctggc aaagaagtac gatgctttcc tggcctctga
     7261 gtctctgatc aagcagattc ctcgtatcct gggccctggc ctcaacaagg caggaaagtt
     7321 tccctctctg ctcacccata atgagaacct gggcaccaag gtggatgagg tgaaatccac
     7381 catcaaattc cagatgaaga aggttctgtg tctggctgtg gccgtgggtc atgtgaagat
     7441 gagtgaggaa gagctggtct acaacatcca cctggctgtc aacttcctgg tgtctctgct
     7501 gaagaagaac tggcaaaacg tcagagctct ttacatcaag agcaccatgg gaaaaccaca
     7561 gcgcctctac tagg
//
