LOCUS       sequence-391381-        4185 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector for endogenously tagging Giardia genes with a triple HA tag 
            (Puromycin selection).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4185)
  AUTHORS   Gourguechon S, Cande WZ
  TITLE     Rapid tagging and integration of genes in Giardia intestinalis.
  JOURNAL   Eukaryot Cell. 2011 Jan;10(1):142-5. doi: 10.1128/EC.00190-10. Epub 
            2010 Nov 29.
  PUBMED    21115739
REFERENCE   2  (bases 1 to 4185)
  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: 
            "10.1128/EC.00190-10"; journalName: "Eukaryot Cell"; date: 
            "2011-01"; volume: "10"; issue: "1"; pages: "142-5"
COMMENT     Sequence Label: pKS_3HA_PAC
FEATURES             Location/Qualifiers
     source          1..4185
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(33..50)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(204..792)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(284..303)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(963..1754,1755..1823))
                     /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     1586..1605
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1824..1928)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      1955..2410
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2042..2061)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2252..2273
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2536..2558
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2550..2567
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2551..2567
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2574..2593
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2574..2592
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2625..2641
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2625..2641
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2675..2691)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2676..2692)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     CDS             2703..2729
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2733..2759
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2763..2789
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3205..3804
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MGTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
                     RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRL
                     AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
                     SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     3704..3724
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     promoter        complement(3948..3966)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3987..4003)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3987..4003)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4000..4022)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4011..4027
                     /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(4035..4041,4042..4059,4060..4065))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4080..4101
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
       61 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
      121 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
      181 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
      241 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
      301 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
      361 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
      421 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
      481 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
      541 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
      601 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
      661 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
      721 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
      781 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
      841 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
      901 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
      961 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     1021 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     1081 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     1141 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     1201 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     1261 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     1321 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     1381 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     1441 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     1501 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     1561 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     1621 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     1681 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     1741 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     1801 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     1861 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     1921 gttccgcgca catttccccg aaaagtgcca cctgacgcgc cctgtagcgg cgcattaagc
     1981 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     2041 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     2101 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     2161 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     2221 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     2281 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     2341 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     2401 cttacaattt ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     2461 cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg
     2521 taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgaa ttgtaatacg
     2581 actcactata gggcgaattg gagctccacc gcggtggcgg ccgctctaga actagtggat
     2641 cccccgggct gcaggaattc gatatcaagc ttatcgatac cgtcgacctc gagcttaagg
     2701 aatatcctta tgacgtccca gactacgcgg agtacccata cgatgtacct gattatgctg
     2761 agtacccata cgacgtacca gattacgctg gatcctgaat cgaattcttt ccgtcgacct
     2821 ttgcatcatc gttgctcggt cgtctgcctc gtttacgacc tgcttgttcc tgggcctcaa
     2881 atggttagtg gttttccttc ttccaacggc gttggtccat cctcggccgc gtgatcaaat
     2941 gccttcgagc gtatgtctga cctgtttgtg aatgacgatc cgtggcttta ctcagattcc
     3001 tgtgtttgtg aataagaaat ccgtgaattt gttggggtgg aacggcgaag tggcctctgc
     3061 tacaataaga actggagagt gtgttcagag gcgaatgcgc aacgaacttt gtcgttacac
     3121 gacgtgtgcg cctgtggtct tcgtgcggtt tccgcgcgga attttgcgcc ttttctaaaa
     3181 aactaaccct aaaacaagaa aatcatgggc accgagtaca agcccacggt gcgcctcgcc
     3241 acccgcgacg acgtcccccg ggccgtacgc accctcgccg ccgcgttcgc cgactacccc
     3301 gccacgcgcc acaccgtcga cccggaccgc cacatcgagc gggtcaccga gctgcaagaa
     3361 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc
     3421 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc
     3481 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc
     3541 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg
     3601 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc
     3661 gagcgcgccg gggtgcctgc cttcctggag acctccgcgc cccgcaacct ccccttctac
     3721 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg
     3781 tgcatgaccc gcaagcccgg tgccctcgag tgagggctga agtgaatatt taccttttcc
     3841 gggttcgacg ggcgtgcttg acgatgcgga agtcgctgtc cgttccaacg gctatctgcc
     3901 ccaggacccc catcccggcg ccgatcagct ggtaccagct tttgttccct ttagtgaggg
     3961 ttaatttcga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     4021 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     4081 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     4141 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggaga
//
