LOCUS       sequence-10323-e        5731 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 5731)
  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 5731)
  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: pCSegfpcherry
FEATURES             Location/Qualifiers
     source          1..5731
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    108..132
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      135..144
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(141..143,144..146,147..857)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(141..143,144..146,147..857)
                     /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(186..207)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(447..466)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     794..815
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    858..923
                     /label=MCS
                     /note="multiple cloning site"
     protein_bind    complement(937..961)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             963..1670
                     /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(1108..1126)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1376..1395
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1570..1593
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     protein_bind    1683..1707
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1720..1740)
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     polyA_signal    1780..1914
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1804..1823)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1858..1877
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2027..2047)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2027..2045)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2066..2082)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2066..2082)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2079..2101)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2090..2106
                     /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(2114..2120,2121..2138,2139..2144))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2159..2180
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2297..2314)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2468..3056)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2548..2567)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3227..4018,4019..4087))
                     /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     3850..3869
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4088..4192)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4218..4673)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4355..4376)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4567..4586
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4747..5731
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagg atcccatcga ttcgaattca aggatcaaca agtttgtaca
      121 aaaaagcagg ctgggccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
      181 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
      241 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
      301 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
      361 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
      421 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
      481 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
      541 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
      601 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
      661 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
      721 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
      781 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
      841 tggacgagct gtacaagtcc ggactcagat ctcgagctca agcttcgaat tctgcagtcg
      901 acggtaccgc gggcccggga tccaccggat ctagacaccc agctttcttg tacaaagtgg
      961 acatggtgag caagggcgag gaggacaaca tggccatcat caaggagttc atgcgcttca
     1021 aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc gagggcgagg
     1081 gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggc ggccccctgc
     1141 ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc tacgtgaagc
     1201 accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc aagtgggagc
     1261 gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg
     1321 acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc gacggccccg
     1381 taatgcagaa gaagaccatg ggctgggagg cctcctccga gcggatgtac cccgaggacg
     1441 gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc cactacgacg
     1501 ccgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc gcctacaacg
     1561 tcaacatcaa gctggacatc acctcccaca acgaggacta caccatcgtg gaacagtacg
     1621 agcgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag agatctcgaa
     1681 tcacaagttt gtacaaaaaa gctgaacgag aaacgtgacc aactttatta tacatagttg
     1741 cactcgagcc tctagaacta tagtgagtcg tattacgtag atccagacat gataagatac
     1801 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     1861 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
     1921 aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaattc
     1981 gcggccgcgg cgccaatgca ttgggcccgg tacccagctt ttgttccctt tagtgagggt
     2041 taattgcgcg cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     2101 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     2161 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     2221 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     2281 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     2341 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     2401 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     2461 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     2521 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     2581 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     2641 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     2701 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     2761 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     2821 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     2881 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     2941 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     3001 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     3061 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     3121 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     3181 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     3241 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     3301 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     3361 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     3421 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     3481 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     3541 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     3601 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     3661 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     3721 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     3781 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     3841 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     3901 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     3961 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     4021 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     4081 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     4141 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     4201 cccgaaaagt gccacctaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt
     4261 ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc
     4321 aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt
     4381 aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact
     4441 acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg
     4501 gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag
     4561 aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac
     4621 gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcccattc
     4681 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     4741 ccagtcgacc atagccaatt caatatggcg tatatggact catgccaatt caatatggtg
     4801 gatctggacc tgtgccaatt caatatggcg tatatggact cgtgccaatt caatatggtg
     4861 gatctggacc ccagccaatt caatatggcg gacttggcac catgccaatt caatatggcg
     4921 gacttggcac tgtgccaact ggggaggggt ctacttggca cggtgccaag tttgaggagg
     4981 ggtcttggcc ctgtgccaag tccgccatat tgaattggca tggtgccaat aatggcggcc
     5041 atattggcta tatgccagga tcaatatata ggcaatatcc aatatggccc tatgccaata
     5101 tggctattgg ccaggttcaa tactatgtat tggccctatg ccatatagta ttccatatat
     5161 gggttttcct attgacgtag atagcccctc ccaatgggcg gtcccatata ccatatatgg
     5221 ggcttcctaa taccgcccat agccactccc ccattgacgt caatggtctc tatatatggt
     5281 ctttcctatt gacgtcatat gggcggtcct attgacgtat atggcgcctc ccccattgac
     5341 gtcaattacg gtaaatggcc cgcctggctc aatgcccatt gacgtcaata ggaccaccca
     5401 ccattgacgt caatgggatg gctcattgcc cattcatatc cgttctcacg ccccctattg
     5461 acgtcaatga cggtaaatgg cccacttggc agtacatcaa tatctattaa tagtaacttg
     5521 gcaagtacat tactattgga aggacgccag ggtacattgg cagtactccc attgacgtca
     5581 atggcggtaa atggcccgcg atggctgcca agtacatccc cattgacgtc aatggggagg
     5641 ggcaatgacg caaatgggcg ttccattgac gtaaatgggc ggtaggcgtg cctaatggga
     5701 ggtctatata agcaatgctc gtttagggaa c
//
