LOCUS       sequence-373847-        6218 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Multisite destination vector for ascidian electroporation of an 
            AttB1/B2 recombined 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 6218)
  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 6218)
  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::RfA
FEATURES             Location/Qualifiers
     source          1..6218
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(332..456)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     polyA_signal    794..928
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(831..850)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     885..904
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(964..983)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(965..983)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1070..1087)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1241..1829)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1321..1340)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2000..2791,2792..2860))
                     /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     2623..2642
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2861..2965)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3033..3051
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3089..3111)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3211..3230
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        3311..3329
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3311..3328
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     3332..3348
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     3333..3349
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3366..3489
                     /gene="mutant version of attR"
                     /label=attR3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(3918..4223)
                     /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"
     CDS             complement(4568..5227)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        complement(join(5281..5287,5288..5305,5306..5311))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(5336..5460)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     5490..5506
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5490..5506
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    5534..5657
                     /gene="mutant version of attR"
                     /label=attR5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        5835..5937
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
ORIGIN
        1 tacacctata aaagagagag ccgttatcgt ctgtttgtgg atgtacagag tgatattatt
       61 gacacgcccg ggcgacggat ggtgatcccc ctggccagtg cacgtctgct gtcagataaa
      121 gtctcccgtg aactttaccc ggtggtgcat atcggggatg aaagctggcg catgatgacc
      181 accgatatgg ccagtgtgcc ggtctccgtt atcggggaag aagtggctga tctcagccac
      241 cgcgaaaatg acatcaaaaa cgccattaac ctgatgttct ggggaatata aatgtcaggc
      301 tcccttatac acagccagtc tgcaggtcga ccatagtgac tggatatgtt gtgttttaca
      361 gtattatgta gtctgttttt tatgcaaaat ctaatttaat atattgatat ttatatcatt
      421 ttacgtttct cgttcagctt tcttgtacaa agtggtgata aacttagata tcggaattcc
      481 agctgagcgc cggtcgctac cattaccagt tggtctggtg tcaaaaataa taataaccgg
      541 gcaggccatg tctgcccgta tttcgcgtaa ggaaatccat tatgtactat ttaaaaaaca
      601 caaacttttg gatgttcggt ttattctttt tcttttactt ttttatcatg ggagcctact
      661 tcccgttttt cccgatttgg ctacatgaca tcaaccatat cagcaaaagt gatacgggta
      721 ttatttttgc cgctatttct ctgttctcgc tattattcca accgctgttt ggtctgcttt
      781 ctgacaaact cggaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
      841 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
      901 catcaatgta tcttatcatg tctggatcga caaagtcaaa gcggccatca gatctgccgg
      961 tctccctata gtgagtcgta ttaatttcga taagccaggt taacctgcat taatgaatcg
     1021 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     1081 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     1141 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     1201 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     1261 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     1321 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     1381 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     1441 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     1501 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     1561 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     1621 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     1681 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     1741 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     1801 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     1861 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     1921 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     1981 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     2041 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     2101 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     2161 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     2221 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     2281 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     2341 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     2401 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     2461 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     2521 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     2581 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     2641 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     2701 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     2761 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     2821 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     2881 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     2941 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag
     3001 aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc
     3061 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
     3121 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
     3181 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
     3241 accatatgga catattgtcg ttagaacgcg gctacaatta atacataacc ttatgtatca
     3301 tacacatacg atttaggtga cactatagaa ctcgaggtcg acggtatcga taagcttgat
     3361 ggggacaagt ttgtataata aagttgaacg agaaacgtaa aatgatataa atatcaatat
     3421 attaaattag attttgcata aaaaacagac tacataatac tgtaaaacac aacatatcca
     3481 gtcactatga atcaactact tagatggtat tagtgacctg tagtcgaccg acagccttcc
     3541 aaatgttctt cgggtgatgc tgccaactta gtcgaccgac agccttccaa atgttcttct
     3601 caaacggaat cgtcgtatcc agcctactcg ctattgtcct caatgccgta ttaaatcata
     3661 aaaagaaata agaaaaagag gtgcgagcct cttttttgtg tgacaaaata aaaacatcta
     3721 cctattcata tacgctagtg tcatagtcct gaaaatcatc tgcatcaaga acaatttcac
     3781 aactcttata cttttctctt acaagtcgtt cggcttcatc tggattttca gcctctatac
     3841 ttactaaacg tgataaagtt tctgtaattt ctactgtatc gacctgcaga ctggctgtgt
     3901 ataagggagc ctgacattta tattccccag aacatcaggt taatggcgtt tttgatgtca
     3961 ttttcgcggt ggctgagatc agccacttct tccccgataa cggagaccgg cacactggcc
     4021 atatcggtgg tcatcatgcg ccagctttca tccccgatat gcaccaccgg gtaaagttca
     4081 cgggagactt tatctgacag cagacgtgca ctggccaggg ggatcaccat ccgtcgcccg
     4141 ggcgtgtcaa taatatcact ctgtacatcc acaaacagac gataacggct ctctctttta
     4201 taggtgtaaa ccttaaactg catttcacca gcccctgttc tcgtcagcaa aagagccgtt
     4261 catttcaata aaccgggcga cctcagccat cccttcctga ttttccgctt tccagcgttc
     4321 ggcacgcaga cgacgggctt cattctgcat ggttgtgctt accagaccgg agatattgac
     4381 atcatatatg ccttgagcaa ctgatagctg tcgctgtcaa ctgtcactgt aatacgctgc
     4441 ttcatagcat acctcttttt gacatacttc gggtatacat atcagtatat attcttatac
     4501 cgcaaaaatc agcgcgcaaa tacgcatact gttatctggc ttttagtaag ccggatccac
     4561 gcgnnnttta cgccccnccc tgccactcat cgcagtactg ttgtaattca ttaagcattc
     4621 tgccgacatg gaagccatca caaacggcat gatgaacctg aatcgccagc ggcatcagca
     4681 ccttgtcgcc ttgcgtataa tatttgccca tggtgaaaac gggggcgaag aagttgtcca
     4741 tattggccac gtttaaatca aaactggtga aactcaccca gggattggct gagacgaaaa
     4801 acatattctc aataaaccct ttagggaaat aggccaggtt ttcaccgtaa cacgccacat
     4861 cttgcgaata tatgtgtaga aactgccgga aatcgtcgtg gtattcactc cagagcgatg
     4921 aaaacgtttc agtttgctca tggaaaacgg tgtaacaagg gtgaacacta tcccatatca
     4981 ccagctcacc gtctttcatt gccatacgga attccggatg agcattcatc aggcgggcaa
     5041 gaatgtgaat aaaggccgga taaaacttgt gcttattttt ctttacggtc tttaaaaagg
     5101 ccgtaatatc cagctgaacg gtctggttat aggtacattg agcaactgac tgaaatgcct
     5161 caaaatgttc tttacgatgc cattgggata tatcaacggt ggtatatcca gtgatttttt
     5221 tctccatttt agcttcctta gctcctgaaa atctcgacgg atcctaactc aaaatccaca
     5281 cattatacga gccggaagca taaagtgtaa agcctggggt gcctaatgcg gccgccatag
     5341 tgactggata tgttgtgttt tacagtatta tgtagtctgt tttttatgca aaatctaatt
     5401 taatatattg atatttatat cattttacgt ttctcgttca gcttttttgt acaaacttgt
     5461 gataaacaaa ggcctattta aatggatcct ctagaactag tggatccccc gggctgcagg
     5521 aattcgatgg ggaccacttt gtatacaaaa gttgaacgag aaacgtaaaa tgatataaat
     5581 atcaatatat taaattagat tttgcataaa aaacagacta cataatactg taaaacacaa
     5641 catatccagt cactatgaat caactactta gatggtatta gtgacctgta gtcgactaag
     5701 ttggcagcat cacccgacgc actttgcgcc gaataaatac ctgtgacgga agatcacttc
     5761 gcagaataaa taaatcctgg tgtccctgtt gataccggga agccctgggc caacttttgg
     5821 cgaaaatgag acgttgatcg gcacgtaaga ggttccaact ttcaccataa tgaaataaga
     5881 tcactaccgg gcgtattttt tgagttatcg agattttcag gagctaagga agctaaaatg
     5941 gagaaaaaaa tcactggata taccaccgtt gatatatccc aatggcatcg taaagaacat
     6001 tttgaggcat ttcagtcagt tgctcaatgt acctataacc agaccgttca gctggatatt
     6061 acggcctttt taaagaccgt aaagaaaaat aagcacaagt tttatccggc ctttattcac
     6121 attcttgccc gcctgatgaa tgctcatccg gaattccgta tggcaatgaa agacggtgag
     6181 ctggtgatat gggatagtgt tcacccttgt tacaccgt
//
