LOCUS       sequence-369045-        4887 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Multisite destination vector for ascidian electroporation of an 
            AttB1/B2 recombined fusion N-ter HA tag-gene of interest under 
            control of an AttB3/B5 recombined reg. sequence..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4887)
  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 4887)
  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-R3-ccdB/CmR-R5::HA-RfA
FEATURES             Location/Qualifiers
     source          1..4887
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     gap             5..79
                     /estimated_length=75
     promoter        complement(join(110..116,117..134,135..140))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(165..289)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(302..328)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     360..376
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     360..376
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    404..527
                     /gene="mutant version of attR"
                     /label=attR5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             808..1467
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(874..893)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             1812..2117
                     /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     1996..2015
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2546..2669)
                     /gene="mutant version of attR"
                     /label=attR3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2686..2702)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2687..2703)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(2706..2724)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2707..2724)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2805..2824)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2924..2946
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2984..3002)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3070..3174
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3175..3243,3244..4035)
                     /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(3393..3412)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4206..4794
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4695..4714
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttacnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn
       61 nnnnnnnnnn nnnnnnnnna tctcgacgga tcctaactca aaatccacac attatacgag
      121 ccggaagcat aaagtgtaaa gcctggggtg cctaatgcgg ccgccatagt gactggatat
      181 gttgtgtttt acagtattat gtagtctgtt ttttatgcaa aatctaattt aatatattga
      241 tatttatatc attttacgtt tctcgttcag cttttttgta caaacttgtg ataaacaaag
      301 gagcgtagtc tggcacgtcg tatgggtatg ccattttttg tccctattta aatggatcct
      361 ctagaactag tggatccccc gggctgcagg aattcgatgg ggaccacttt gtatacaaaa
      421 gttgaacgag aaacgtaaaa tgatataaat atcaatatat taaattagat tttgcataaa
      481 aaacagacta cataatactg taaaacacaa catatccagt cactatgaat caactactta
      541 gatggtatta gtgacctgta gtcgactaag ttggcagcat cacccgacgc actttgcgcc
      601 gaataaatac ctgtgacgga agatcacttc gcagaataaa taaatcctgg tgtccctgtt
      661 gataccggga agccctgggc caacttttgg cgaaaatgag acgttgatcg gcacgtaaga
      721 ggttccaact ttcaccataa tgaaataaga tcactaccgg gcgtatkttw tgakyyrtcg
      781 agattttcag gagctaagga agctaaaatg gagaaaaaaa tcactggata taccaccgtt
      841 gatatatccc aatggcatcg taaagaacat tttgaggcat ttcagtcagt tgctcaatgt
      901 acctataacc agaccgttca gctggatatt acggcctttt taaagaccgt aaagaaaaat
      961 aagcacaagt tttatccggc ctttattcac attcttgccc gcctgatgaa tgctcatccg
     1021 gaattccgta tggcaatgaa agacggtgag ctggtgatat gggatagtgt tcacccttgt
     1081 tacaccgttt tccatgagca aactgaaacg ttttcatcgc tctggagtga ataccacgac
     1141 gatttccggc agtttctaca catatattcg caagatgtgg cgtgttacgg tgaaaacctg
     1201 gcctatttcc ctaaagggtt tattgagaat atgtttttcg tctcagccaa tccctgggtg
     1261 agtttcacca gttttgattt aaacgtggcc aatatggaca acttcttcgc ccccgttttc
     1321 accatgggca aatattatac gcaaggcgac aaggtgctga tgccgctggc gattcaggtt
     1381 catcatgccg tttgtgatgg cttccatgtc ggcagaatgc ttaatgaatt acaacagtac
     1441 tgcgatgagt ggcagggngg ggcgtaaacg cngnntggat ccggcttact aaaagccaga
     1501 taacagtatg cgtatttgcg cgctgatttt tgcggtataa gaatatatac tgatatgtat
     1561 acccgaagta tgtcaaaaag aggtatgcta tgaagcagcg tattacagtg acagttgaca
     1621 gcgacagcta tcagttgctc aaggcatata tgatgtcaat atctccggtc tggtaagcac
     1681 aaccatgcag aatgaagccc gtcgtctgcg tgccgaacgc tggaaagcgg aaaatcagga
     1741 agggatggct gaggtcgccc ggtttattga aatgaacggc tcttttgctg acgagaacag
     1801 gggctggtga aatgcagttt aaggtttaca cctataaaag agagagccgt tatcgtctgt
     1861 ttgtggatgt acagagtgat attattgaca cgcccgggcg acggatggtg atccccctgg
     1921 ccagtgcacg tctgctgtca gataaagtct cccgtgaact ttacccggtg gtgcatatcg
     1981 gggatgaaag ctggcgcatg atgaccaccg atatggccag tgtgccggtc tccgttatcg
     2041 gggaagaagt ggctgatctc agccaccgcg aaaatgacat caaaaacgcc attaacctga
     2101 tgttctgggg aatataaatg tcaggctccc ttatacacag ccagtctgca ggtcgataca
     2161 gtagaaatta cagaaacttt atcacgttta gtaagtatag aggctgaaaa tccagatgaa
     2221 gccgaacgac ttgtaagaga aaagtataag agttgtgaaa ttgttcttga tgcagatgat
     2281 tttcaggact atgacactag cgtatatgaa taggtagatg tttttatttt gtcacacaaa
     2341 aaagaggctc gcacctcttt ttcttatttc tttttatgat ttaatacggc attgaggaca
     2401 atagcgagta ggctggatac gacgattccg tttgagaaga acatttggaa ggctgtcggt
     2461 cgactaagtt ggcagcatca cccgaagaac atttggaagg ctgtcggtcg actacaggtc
     2521 actaatacca tctaagtagt tgattcatag tgactggata tgttgtgttt tacagtatta
     2581 tgtagtctgt tttttatgca aaatctaatt taatatattg atatttatat cattttacgt
     2641 ttctcgttca actttattat acaaacttgt ccccatcaag cttatcgata ccgtcgacct
     2701 cgagttctat agtgtcacct aaatcgtatg tgtatgatac ataaggttat gtattaattg
     2761 tagccgcgtt ctaacgacaa tatgtccata tggtgcactc tcagtacaat ctgctctgat
     2821 gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     2881 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     2941 cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta
     3001 tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg
     3061 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     3121 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     3181 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     3241 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     3301 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     3361 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     3421 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     3481 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     3541 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     3601 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     3661 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     3721 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     3781 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     3841 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     3901 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     3961 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     4021 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     4081 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     4141 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     4201 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     4261 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     4321 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     4381 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     4441 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     4501 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     4561 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     4621 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     4681 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     4741 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     4801 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     4861 cctgcgttat cccctgattc tgtggat
//
