LOCUS       sequence-328789-        3440 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Template plasmid for generating C-terminal 6-His epitope tag fusions
            to bacterial genes by ""Lambda red"" recombineering..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3440)
  AUTHORS   Uzzau S, Figueroa-Bossi N, Rubino S, Bossi L
  TITLE     Epitope tagging of chromosomal genes in Salmonella.
  JOURNAL   Proc Natl Acad Sci U S A. 2001 Dec 18;98(26):15264-9. doi: 
            10.1073/pnas.261348198. Epub 2001 Dec 11.
  PUBMED    11742086
REFERENCE   2  (bases 1 to 3440)
  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"; doi: 
            "10.1073/pnas.261348198"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2001-12-18- 18"; volume: "98"; issue: "26"; pages: "15264-9"
COMMENT     Sequence Label: pSUB7
FEATURES             Location/Qualifiers
     source          1..3440
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        28..132
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(133..201,202..993)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(351..370)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     primer_bind     complement(1068..1087)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1228..1250
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1389..1408
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     rep_origin      1492..1848
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     CDS             1913..1930
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    complement(1956..2003)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             2364..3158
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2418..2437)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3028..3047
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    complement(3349..3382)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
ORIGIN
        1 gcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
       61 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
      121 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
      181 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
      241 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
      301 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
      361 gcggtattat cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat acactattct
      421 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
      481 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
      541 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
      601 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
      661 gacaccacga tgcctgcagc aatggcaaca acgttgcgca aactattaac tggcgaacta
      721 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
      781 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
      841 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
      901 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
      961 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     1021 ctttagattg atttaaaact tcatttttaa ttttggtgca ctctcagtac aatctgctct
     1081 gatgccgcat agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg
     1141 cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     1201 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     1261 catcaccgaa acgcgcgagg cagcaaggag atggcgccca acagtccccc ggccacgggg
     1321 cctgccacca tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct
     1381 tccccatcgg tgatgtcggc gatataggcg ccagcaaccg cacctgtggc gccggtgatg
     1441 ccggccacga tgcgtccggc gtagaggatc cagcagttca acctgttgat agtacgtact
     1501 aagctctcat gtttcacgta ctaagctctc atgtttaacg tactaagctc tcatgtttaa
     1561 cgaactaaac cctcatggct aacgtactaa gctctcatgg ctaacgtact aagctctcat
     1621 gtttcacgta ctaagctctc atgtttgaac aataaaatta atataaatca gcaacttaaa
     1681 tagcctctaa ggttttaagt tttataagaa aaaaaagaat atataaggct tttaaagctt
     1741 ttaaggttta acggttgtgg acaacaagcc agggatgtaa cgcactgaga agcccttaga
     1801 gcctctcaaa gcaattttca gtgacacagg aacacttaac ggctgacatg gccttagaag
     1861 gcgttttaga agccctaatg gatggagtca aaacaaacta gcgatcgaat tccaccacca
     1921 tcatcaccat tagtaatgta ggctggagct gcttcgaagt tcctatactt tctagagaat
     1981 aggaacttcg gaataggaac ttcaagatcc cctcacgctg ccgcaagcac tcagggcgca
     2041 agggctgcta aaggaagcgg aacacgtaga aagccagtcc gcagaaacgg tgctgacccc
     2101 ggatgaatgt cagctactgg gctatctgga caagggaaaa cgcaagcgca aagagaaagc
     2161 aggtagcttg cagtgggctt acatggcgat agctagactg ggcggtttta tggacagcaa
     2221 gcgaaccgga attgccagct ggggcgccct ctggtaaggt tgggaagccc tgcaaagtaa
     2281 actggatggc tttcttgccg ccaaggatct gatggcgcag gggatcaaga tctgatcaag
     2341 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     2401 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     2461 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     2521 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     2581 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     2641 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     2701 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     2761 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     2821 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     2881 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     2941 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     3001 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     3061 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     3121 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     3181 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     3241 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     3301 ggatctcatg ctggagttct tcgcccaccc cagcttcaaa agcgctctga agttcctata
     3361 ctttctagag aataggaact tcggaatagg aactaaggag gatattcata tgcggaattc
     3421 ccgggagagc tcgatgatct
//
