LOCUS       sequence-373585-        4872 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Destination vector for the expression of a fusion of an N-ter 
            protein of interest with an HA tag into embryos, by mRNA 
            microinjection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4872)
  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 4872)
  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-Spe3-RfA-HA
FEATURES             Location/Qualifiers
     source          1..4872
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(80..99)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     290..311
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     575..597
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     589..606
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     590..606
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     613..632
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        613..631
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     3'UTR           complement(676..801)
                     /label=Xenopus globin 3'-UTR
                     /note="3'-UTR of the major beta-globin gene of Xenopus 
                     laevis"
     CDS             complement(833..859)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    873..997
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(1038..1343)
                     /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     complement(1140..1159)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1685..2344)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2259..2278
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(2398..2404,2405..2422,2423..2428))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(2453..2577)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     5'UTR           complement(2606..2648)
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     promoter        complement(2673..2691)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2712..2728)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2712..2728)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2725..2747)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2736..2752
                     /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(2760..2766,2767..2784,2785..2790))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2805..2826
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2943..2960)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3114..3702)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3194..3213)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3873..4664,4665..4733))
                     /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     4496..4515
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4734..4838)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      join(4865..4872,1..448)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
       61 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
      121 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
      181 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
      241 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
      301 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
      361 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
      421 tttaacaaaa tattaacgct tacaatttgc cattcgccat tcaggctgcg caactgttgg
      481 gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct
      541 gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg
      601 gccagtgaat tgtaatacga ctcactatag ggcgaattgg gtaccggccg acttggccct
      661 gcaggggggg gggnngtgaa gaaactttct ttttattagg agcagatacg aatggctaca
      721 ttttggggga caacattttg taaagtgtaa gttggtatta tgtagcttag agactccatt
      781 cgggtgttct tgaggctggt ttagtggtaa ccagatccgc ggccgcgatt caagcgtagt
      841 ctggcacgtc gtatgggtaa tctaagttta tcaccacttt gtacaagaaa gctgaacgag
      901 aaacgtaaaa tgatataaat atcaatatat taaattagat tttgcataaa aaacagacta
      961 cataatactg taaaacacaa catatccagt cactatggtc gacctgcaga ctggctgtgt
     1021 ataagggagc ctgacattta tattccccag aacatcaggt taatggcgtt tttgatgtca
     1081 ttttcgcggt ggctgagatc agccacttct tccccgataa cggagaccgg cacactggcc
     1141 atatcggtgg tcatcatgcg ccagctttca tccccgatat gcaccaccgg gtaaagttca
     1201 cgggagactt tatctgacag cagacgtgca ctggccaggg ggatcaccat ccgtcgcccg
     1261 ggcgtgtcaa taatatcact ctgtacatcc acaaacagac gataacggct ctctctttta
     1321 taggtgtaaa ccttaaactg catttcacca gcccctgttc tcgtcagcaa aagagccgtt
     1381 catttcaata aaccgggcga cctcagccat cccttcctga ttttccgctt tccagcgttc
     1441 ggcacgcaga cgacgggctt cattctgcat ggttgtgctt accagaccgg agatattgac
     1501 atcatatatg ccttgagcaa ctgatagctg tcgctgtcaa ctgtcactgt aatacgctgc
     1561 ttcatagcat acctcttttt gacatacttc gggtatacat atcagtatat attcttatac
     1621 cgcaaaaatc agcgcgcaaa tacgcatact gttatctggc ttttagtaag ccggatccac
     1681 gcgtttacgc cccgccctgc cactcatcgc agtactgttg taattcatta agcattctgc
     1741 cgacatggaa gccatcacaa acggcatgat gaacctgaat cgccagcggc atcagcacct
     1801 tgtcgccttg cgtataatat ttgcccatgg tgaaaacggg ggcgaagaag ttgtccatat
     1861 tggccacgtt taaatcaaaa ctggtgaaac tcacccaggg attggctgag acgaaaaaca
     1921 tattctcaat aaacccttta gggaaatagg ccaggttttc accgtaacac gccacatctt
     1981 gcgaatatat gtgtagaaac tgccggaaat cgtcgtggta ttcactccag agcgatgaaa
     2041 acgtttcagt ttgctcatgg aaaacggtgt aacaagggtg aacactatcc catatcacca
     2101 gctcaccgtc tttcattgcc atacggaatt ccggatgagc attcatcagg cgggcaagaa
     2161 tgtgaataaa ggccggataa aacttgtgct tatttttctt tacggtcttt aaaaaggccg
     2221 taatatccag ctgaacggtc tggttatagg tacattgagc aactgactga aatgcctcaa
     2281 aatgttcttt acgatgccat tgggatatat caacggtggt atatccagtg atttttttct
     2341 ccattttagc ttccttagct cctgaaaatc tcgacggatc ctaactcaaa atccacacat
     2401 tatacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgcggcc gccatagtga
     2461 ctggatatgt tgtgttttac agtattatgt agtctgtttt ttatgcaaaa tctaatttaa
     2521 tatattgata tttatatcat tttacgtttc tcgttcagct tttttgtaca aacttgtgat
     2581 aaacaaaggc ctgaattcag atctgccaaa gttgagcgtt tattctgagc ttctgcaaaa
     2641 agaacaagca agcttagctc cagcttttgt tccctttagt gagggttaat ttcgagcttg
     2701 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     2761 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     2821 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     2881 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     2941 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     3001 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     3061 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     3121 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     3181 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     3241 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     3301 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     3361 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     3421 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     3481 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     3541 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     3601 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     3661 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     3721 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     3781 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     3841 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     3901 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     3961 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4021 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4081 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     4141 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     4201 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     4261 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     4321 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     4381 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     4441 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     4501 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     4561 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     4621 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     4681 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     4741 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     4801 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     4861 cctgacgcgc cc
//
