LOCUS       sequence-373584-        4886 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Destination vector for the expression of a fusion of N-ter HA tag 
            and a protein of interest into embryos, by mRNA microinjection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4886)
  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 4886)
  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-HA-RfA
FEATURES             Location/Qualifiers
     source          1..4886
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      12..467
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(99..118)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     309..330
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     594..616
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     608..625
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     609..625
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     632..651
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        632..650
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     3'UTR           complement(698..823)
                     /label=Xenopus globin 3'-UTR
                     /note="3'-UTR of the major beta-globin gene of Xenopus 
                     laevis"
     protein_bind    865..989
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(1030..1335)
                     /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(1132..1151)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1677..2336)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2251..2270
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(2390..2396,2397..2414,2415..2420))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(2445..2569)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(2582..2608)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     5'UTR           complement(2639..2681)
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     promoter        complement(2706..2724)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2745..2761)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2745..2761)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2758..2780)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2769..2785
                     /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(2793..2799,2800..2817,2818..2823))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2838..2859
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2976..2993)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3147..3735)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3227..3246)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3906..4697,4698..4766))
                     /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     4529..4548
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4767..4871)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 agtgccacct gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
       61 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
      121 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
      181 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
      241 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
      301 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
      361 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
      421 acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttgcc attcgccatt
      481 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
      541 ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc
      601 acgacgttgt aaaacgacgg ccagtgaatt gtaatacgac tcactatagg gcgaattggg
      661 taccggccga cttggccctg cagggggggg gnnnaatgtg aagaaacttt ctttttatta
      721 ggagcagata cgaatggcta cattttgggg gacaacattt tgtaaagtgt aagttggtat
      781 tatgtagctt agagactcca ttcgggtgtt cttgaggctg gtttagtggt aaccagatcc
      841 gcggccgcga tatctaagtt tatcaccact ttgtacaaga aagctgaacg agaaacgtaa
      901 aatgatataa atatcaatat attaaattag attttgcata aaaaacagac tacataatac
      961 tgtaaaacac aacatatcca gtcactatgg tcgacctgca gactggctgt gtataaggga
     1021 gcctgacatt tatattcccc agaacatcag gttaatggcg tttttgatgt cattttcgcg
     1081 gtggctgaga tcagccactt cttccccgat aacggagacc ggcacactgg ccatatcggt
     1141 ggtcatcatg cgccagcttt catccccgat atgcaccacc gggtaaagtt cacgggagac
     1201 tttatctgac agcagacgtg cactggccag ggggatcacc atccgtcgcc cgggcgtgtc
     1261 aataatatca ctctgtacat ccacaaacag acgataacgg ctctctcttt tataggtgta
     1321 aaccttaaac tgcatttcac cagcccctgt tctcgtcagc aaaagagccg ttcatttcaa
     1381 taaaccgggc gacctcagcc atcccttcct gattttccgc tttccagcgt tcggcacgca
     1441 gacgacgggc ttcattctgc atggttgtgc ttaccagacc ggagatattg acatcatata
     1501 tgccttgagc aactgatagc tgtcgctgtc aactgtcact gtaatacgct gcttcatagc
     1561 atacctcttt ttgacatact tcgggtatac atatcagtat atattcttat accgcaaaaa
     1621 tcagcgcgca aatacgcata ctgttatctg gcttttagta agccggatcc acgcgtttac
     1681 gccccgccct gccactcatc gcagtactgt tgtaattcat taagcattct gccgacatgg
     1741 aagccatcac aaacggcatg atgaacctga atcgccagcg gcatcagcac cttgtcgcct
     1801 tgcgtataat atttgcccat ggtgaaaacg ggggcgaaga agttgtccat attggccacg
     1861 tttaaatcaa aactggtgaa actcacccag ggattggctg agacgaaaaa catattctca
     1921 ataaaccctt tagggaaata ggccaggttt tcaccgtaac acgccacatc ttgcgaatat
     1981 atgtgtagaa actgccggaa atcgtcgtgg tattcactcc agagcgatga aaacgtttca
     2041 gtttgctcat ggaaaacggt gtaacaaggg tgaacactat cccatatcac cagctcaccg
     2101 tctttcattg ccatacggaa ttccggatga gcattcatca ggcgggcaag aatgtgaata
     2161 aaggccggat aaaacttgtg cttatttttc tttacggtct ttaaaaaggc cgtaatatcc
     2221 agctgaacgg tctggttata ggtacattga gcaactgact gaaatgcctc aaaatgttct
     2281 ttacgatgcc attgggatat atcaacggtg gtatatccag tgattttttt ctccatttta
     2341 gcttccttag ctcctgaaaa tctcgacgga tcctaactca aaatccacac attatacgag
     2401 ccggaagcat aaagtgtaaa gcctggggtg cctaatgcgg ccgccatagt gactggatat
     2461 gttgtgtttt acagtattat gtagtctgtt ttttatgcaa aatctaattt aatatattga
     2521 tatttatatc attttacgtt tctcgttcag cttttttgta caaacttgtg ataaacaaag
     2581 gagcgtagtc tggcacgtcg tatgggtatg ccattttttg tccctgaatt cagatctgcc
     2641 aaagttgagc gtttattctg agcttctgca aaaagaacaa gcaagcttag ctccagcttt
     2701 tgttcccttt agtgagggtt aatttcgagc ttggcgtaat catggtcata gctgtttcct
     2761 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     2821 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     2881 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     2941 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     3001 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     3061 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     3121 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     3181 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     3241 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     3301 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     3361 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     3421 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     3481 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     3541 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     3601 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     3661 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     3721 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     3781 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     3841 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     3901 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     3961 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     4021 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     4081 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     4141 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     4201 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     4261 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     4321 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     4381 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     4441 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     4501 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     4561 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     4621 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     4681 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     4741 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     4801 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     4861 ggggttccgc gcacatttcc ccgaaa
//
