LOCUS       sequence-373699-        5370 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Destination vector for ascidian transgenesis by electroporation of a
            Gateway-cloned fusion of a gene of interest with a C-ter eCFP under 
            control of a reg. sequence in the Swa1 restriction site..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5370)
  AUTHORS   Roure A, Rothbacher U, Robin F, Kalmar E, Ferone G, Lamy C, Missero 
            C, Mueller F, Lemaire P
  TITLE     A multicassette Gateway vector set for high throughput and 
            comparative analyses in ciona and vertebrate embryos.
  JOURNAL   PLoS One. 2007. 2(9):e916.
  PUBMED    17878951
REFERENCE   2  (bases 1 to 5370)
  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: "PLoS One. 
            2007. 2(9):e916."
COMMENT     Sequence Label: pDEST-pSP172BSSPE-Swa1::RfA-ECFP
FEATURES             Location/Qualifiers
     source          1..5370
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    83..207
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(232..237,238..255,256..262)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             316..975
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(382..401)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             1317..1622
                     /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     1501..1520
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(1663..1787)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             join(1801..1803,1804..1806,1807..2520)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1846..1867)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2107..2126)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2454..2475
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2845..2979
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2882..2901)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2936..2955
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3015..3034)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3016..3034)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3121..3138)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3292..3880)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3372..3391)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4051..4842,4843..4911))
                     /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     4674..4693
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4912..5016)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5084..5102
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5140..5162)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5262..5281
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(5362..5370,1..10)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(5362..5370,1..9)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 acactataga actcgagcag ctgaagcttg catgcctgca ggtcgactct agaggatcca
       61 tttaaatagg cctttgttta tcacaagttt gtacaaaaaa gctgaacgag aaacgtaaaa
      121 tgatataaat atcaatatat taaattagat tttgcataaa aaacagacta cataatactg
      181 taaaacacaa catatccagt cactatggcg gccgcattag gcaccccagg ctttacactt
      241 tatgcttccg gctcgtataa tgtgtggatt ttgagttagg atccgtcgag attttcagga
      301 gctaaggaag ctaaaatgga gaaaaaaatc actggatata ccaccgttga tatatcccaa
      361 tggcatcgta aagaacattt tgaggcattt cagtcagttg ctcaatgtac ctataaccag
      421 accgttcagc tggatattac ggccttttta aagaccgtaa agaaaaataa gcacaagttt
      481 tatccggcct ttattcacat tcttgcccgc ctgatgaatg ctcatccgga attccgtatg
      541 gcaatgaaag acggtgagct ggtgatatgg gatagtgttc acccttgtta caccgttttc
      601 catgagcaaa ctgaaacgtt ttcatcgctc tggagtgaat accacgacga tttccggcag
      661 tttctacaca tatattcgca agatgtggcg tgttacggtg aaaacctggc ctatttccct
      721 aaagggttta ttgagaatat gtttttcgtc tcagccaatc cctgggtgag tttcaccagt
      781 tttgatttaa acgtggccaa tatggacaac ttcttcgccc ccgttttcac catgggcaaa
      841 tattatacgc aaggcgacaa ggtgctgatg ccgctggcga ttcaggttca tcatgccgtt
      901 tgtgatggct tccatgtcgg cagaatgctt aatgaattac aacagtactg cgatgagtgg
      961 cagggcgggg cgtaaacgcg tggatccggc ttactaaaag ccagataaca gtatgcgtat
     1021 ttgcgcgctg atttttgcgg tataagaata tatactgata tgtatacccg aagtatgtca
     1081 aaaagaggta tgctatgaag cagcgtatta cagtgacagt tgacagcgac agctatcagt
     1141 tgctcaaggc atatatgatg tcaatatctc cggtctggta agcacaacca tgcagaatga
     1201 agcccgtcgt ctgcgtgccg aacgctggaa agcggaaaat caggaaggga tggctgaggt
     1261 cgcccggttt attgaaatga acggctcttt tgctgacgag aacaggggct ggtgaaatgc
     1321 agtttaaggt ttacacctat aaaagagaga gccgttatcg tctgtttgtg gatgtacaga
     1381 gtgatattat tgacacgccc gggcgacgga tggtgatccc cctggccagt gcacgtctgc
     1441 tgtcagataa agtctcccgt gaactttacc cggtggtgca tatcggggat gaaagctggc
     1501 gcatgatgac caccgatatg gccagtgtgc cggtctccgt tatcggggaa gaagtggctg
     1561 atctcagcca ccgcgaaaat gacatcaaaa acgccattaa cctgatgttc tggggaatat
     1621 aaatgtcagg ctcccttata cacagccagt ctgcaggtcg accatagtga ctggatatgt
     1681 tgtgttttac agtattatgt agtctgtttt ttatgcaaaa tctaatttaa tatattgata
     1741 tttatatcat tttacgtttc tcgttcagct ttcttgtaca aagtggtgat aaacttagat
     1801 atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     1861 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
     1921 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     1981 ctcgtgacca ccctgacctg gggcgtgcag tgcttcagcc gctaccccga ccacatgaag
     2041 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     2101 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     2161 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     2221 aagctggagt acaactacat cagccacaac gtctatatca ccgccgacaa gcagaagaac
     2281 ggcatcaagg ccaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
     2341 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     2401 tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     2461 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa
     2521 atcggaattc cagctgagcg ccggtcgcta ccattaccag ttggtctggt gtcaaaaata
     2581 ataataaccg ggcaggccat gtctgcccgt atttcgcgta aggaaatcca ttatgtacta
     2641 tttaaaaaac acaaactttt ggatgttcgg tttattcttt ttcttttact tttttatcat
     2701 gggagcctac ttcccgtttt tcccgatttg gctacatgac atcaaccata tcagcaaaag
     2761 tgatacgggt attatttttg ccgctatttc tctgttctcg ctattattcc aaccgctgtt
     2821 tggtctgctt tctgacaaac tcggaacttg tttattgcag cttataatgg ttacaaataa
     2881 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     2941 ttgtccaaac tcatcaatgt atcttatcat gtctggatcg acaaagtcaa agcggccatc
     3001 agatctgccg gtctccctat agtgagtcgt attaatttcg ataagccagg ttaacctgca
     3061 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     3121 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     3181 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     3241 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     3301 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     3361 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     3421 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     3481 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     3541 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     3601 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     3661 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     3721 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     3781 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     3841 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     3901 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     3961 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     4021 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     4081 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     4141 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     4201 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     4261 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     4321 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     4381 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     4441 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     4501 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     4561 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     4621 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     4681 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     4741 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     4801 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     4861 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     4921 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     4981 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     5041 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
     5101 gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     5161 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     5221 gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt
     5281 actgagagtg caccatatgg acatattgtc gttagaacgc ggctacaatt aatacataac
     5341 cttatgtatc atacacatac gatttaggtg
//
