LOCUS       sequence-70499-e        2745 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Contributes the coding sequence of a V5-6xHis epitope tags cassette 
            as the 5'-module during MultiSite Gateway-cloning of chimeric cDNAs 
            encoding three-part fusion proteins..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2745)
  AUTHORS   Buj R, Iglesias N, Planas AM, Santalucia T
  TITLE     A plasmid toolkit for cloning chimeric cDNAs encoding customized 
            fusion proteins into any Gateway destination expression vector.
  JOURNAL   BMC Mol Biol. 2013 Aug 20;14(1):18. doi: 10.1186/1471-2199-14-18.
  PUBMED    23957834
REFERENCE   2  (bases 1 to 2745)
  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: "BMC Mol 
            Biol."; date: "2013-08-20"; volume: "14(1):18. doi"; pages: " 
            10.1186/1471-2199-14-18"
COMMENT     Sequence Label: pDONR P4-P1R-V5-6xHis
FEATURES             Location/Qualifiers
     source          1..2745
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    593..688
                     /gene="mutant version of attL"
                     /label=attL4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      691..700
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             721..762
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             772..789
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    790..914
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(986..1005)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(987..1005)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1010..1026)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1010..1026)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1128..1147)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1139..1948
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1206..1225)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2095..2683
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2584..2603
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccgc gttaacgcta ccatggagct ccaaataatg
      601 attttatttt gactgatagt gacctgttcg ttgcaacaaa ttgataagca atgctttttt
      661 ataatgccaa ctttgtatag aaaagttgcc gccaccatgg gtaccgagct cggatccgaa
      721 ggtaagccta tccctaaccc tctcctcggt ctcgattcta cgcgtaccgg tcatcatcac
      781 catcaccata caagtttgta caaaaaagtt gaacgagaaa cgtaaaatga tataaatatc
      841 aatatattaa attagatttt gcataaaaaa cagactacat aatactgtaa aacacaacat
      901 atgcagtcac tatgaaccaa ctacttagat ggtattagtg acctgtagaa ttcgagctct
      961 agagctgcag ggcggccgcg atatccccta tagtgagtcg tattacatgg tcatagctgt
     1021 ttcctggcag ctctggcccg tgtctcaaaa tctctgatgt tacattgcac aagataaaaa
     1081 tatatcatca tgaacaataa aactgtctgc ttacataaac agtaatacaa ggggtgttat
     1141 gagccatatt caacgggaaa cgtcgaggcc gcgattaaat tccaacatgg atgctgattt
     1201 atatgggtat aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa tctatcgctt
     1261 gtatgggaag cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa
     1321 tgatgttaca gatgagatgg tcagactaaa ctggctgacg gaatttatgc ctcttccgac
     1381 catcaagcat tttatccgta ctcctgatga tgcatggtta ctcaccactg cgatccccgg
     1441 aaaaacagca ttccaggtat tagaagaata tcctgattca ggtgaaaata ttgttgatgc
     1501 gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc cttttaacag
     1561 cgatcgcgta tttcgtctcg ctcaggcgca atcacgaatg aataacggtt tggttgatgc
     1621 gagtgatttt gatgacgagc gtaatggctg gcctgttgaa caagtctgga aagaaatgca
     1681 taaacttttg ccattctcac cggattcagt cgtcactcat ggtgatttct cacttgataa
     1741 ccttattttt gacgagggga aattaatagg ttgtattgat gttggacgag tcggaatcgc
     1801 agaccgatac caggatcttg ccatcctatg gaactgcctc ggtgagtttt ctccttcatt
     1861 acagaaacgg ctttttcaaa aatatggtat tgataatcct gatatgaata aattgcagtt
     1921 tcatttgatg ctcgatgagt ttttctaatc agaattggtt aattggttgt aacactggca
     1981 gagcattacg ctgacttgac gggacggcgc aagctcatga ccaaaatccc ttaacgtgag
     2041 ttacgcgtcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     2101 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     2161 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     2221 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     2281 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     2341 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     2401 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     2461 cgaactgaga tacctacagc gtgagcattg agaaagcgcc acgcttcccg aagggagaaa
     2521 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     2581 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     2641 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     2701 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgtt
//
