LOCUS       sequence-433103-        8065 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Platform for DiCre inducible expression of a gene of interest in 
            Leishmania major - requires Leishmania strain expressing DiCre 
            recombinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8065)
  AUTHORS   Santos RERS, Silva GLA, Santos EV, Duncan SM, Mottram JC, Damasceno 
            JD, Tosi LRO
  TITLE     A DiCre recombinase-based system for inducible expression in 
            Leishmania major.
  JOURNAL   Mol Biochem Parasitol. 2017 Sep;216:45-48. doi: 
            10.1016/j.molbiopara.2017.06.006. Epub 2017 Jun 16.
  PUBMED    28629935
REFERENCE   2  (bases 1 to 8065)
  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: "Mol Biochem
            Parasitol."; date: "2017-09"; volume: "216:45-48. doi"; pages: " 
            10.1016/j.molbiopara.2017.06.006. Epub 2017 Jun 16"
COMMENT     Sequence Label: pGL6000
FEATURES             Location/Qualifiers
     source          1..8065
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(1..589)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(81..100)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(760..1551,1552..1620))
                     /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     1383..1402
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1621..1725)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1793..1811
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1849..1871)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1971..1990
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2179..2201
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2193..2210
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2194..2210
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2679..2712)
                     /label=lox66
                     /bound_moiety="Cre recombinase"
                     /note="Right element (RE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(2783..2809)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2816..2842)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2849..2875)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2882..2908)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2915..2941)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2948..2974)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    3025..3058
                     /label=lox71
                     /bound_moiety="Cre recombinase"
                     /note="Left element (LE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(3483..3494)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             4751..5350
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4751..4770)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5247..5267
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(7664..7680)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7664..7680)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7677..7699)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7688..7704
                     /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(7712..7718,7719..7736,7737..7742))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7757..7778
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7895..7912)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
       61 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
      121 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
      181 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
      241 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
      301 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
      361 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
      421 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
      481 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
      541 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
      601 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
      661 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
      721 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
      781 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
      841 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
      901 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
      961 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     1021 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     1081 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     1141 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     1201 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     1261 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     1321 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     1381 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     1441 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     1501 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     1561 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     1621 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     1681 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     1741 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     1801 tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     1861 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     1921 tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga
     1981 gcagattgta ctgagagtgc acccggtgtg aaataccgca cagatgcgta aggagaaaat
     2041 accgcatcag gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc
     2101 gggcctcttc gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt
     2161 gggtaacgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt gaatgatcct
     2221 taattaatga cgaacaactg ccctatcagc tggtgatggc cgtgtagtgg actgccatgg
     2281 cgttgacggg agcgggggat tagggttcga ttccggagag ggagcctgag aaatagctac
     2341 cacttctacg gagggcagca ggcgcgcaaa ttgcccaatg tcaaaacaaa acgatgaggc
     2401 agcgaaaaga aatagagttg tcagtccatt tggattgtca tttcaatggg ggatatttaa
     2461 acccatccaa tatcgagtaa caattgcccg ctttccattt cgtcaccttc cgcctctctc
     2521 tctctctctc tcaccatcta cgcgtgcaca tcatcaactg tctcttgtcg gtgctcacca
     2581 ccctcaacca cccctcactt tcaaggcttc ccgaacgcac acaaaaggcg tgaaaaccgc
     2641 tcgcggtgtg ttgagccgtc caccgtagcc tcgaggtgat aacttcgtat agcatacatt
     2701 atacgaacgg tactcctatc tagaactagt atcgatgagc tctcgcgagg taccctagcg
     2761 tggatctgga tccttacctt gaggcataat caggaacgtc gtaggggtaa cttgaggcat
     2821 aatcaggaac gtcgtagggg taacttgagg cataatcagg aacgtcgtag gggtaacttg
     2881 aggcataatc aggaacgtcg taggggtaac ttgaggcata atcaggaacg tcgtaggggt
     2941 aacttgaggc ataatcagga acgtcgtagg ggtatctaca ccgccctgca ggaatcatat
     3001 gaaatagatc ctaatgctag cgtgataact tcgtataatg tatgctatac gaacggtact
     3061 cgcggccgcg gcgcgcctta agcaagagca tagccttact tgcgggcttc ctcaacagta
     3121 caccccctat tcgacagtgc ttctgcgaga aatcgtacat ctgcgctcag gttgcaaatt
     3181 cgcaaacgcc caacacacga acattctctg atttattcta tctgcctcgg tacatgcttt
     3241 tcttccttct gtgcggtgtg aagcatattt gcagtggatt atgctttcgc ttagccgtgt
     3301 ttttgtttct cccactacta ctgctgctac tgttcttttc gtgttctagc ccttcgaggg
     3361 gccctagtag tgatcccgcc cgcacccgcg cgcgctcagg ccgcgtgtcg tcgtgctctt
     3421 tccatttttt ttttgtgtgt ggtggtgatt tgagctgctc tccgttgtgt gctggggacc
     3481 ctccttccct cgatctcctc gtgcggggtc tccgacgccg cagacgcggt gcgtgcgagc
     3541 gtgcacgtgg ctgtcgccgt ggctgtagtt gcgagcggag agagagagag agggagggag
     3601 ggagggcaga gggcagagga catgcgggtg ggaacgtgca ccggcctcgt ctcacgcagc
     3661 tggagcccac gaatgccacc accacaccct tcttcccccc ccctccctct tcttcccacg
     3721 gcggcgacga cgacggggct cagctcacgc ttctgtaggg tattatatta aagcacatgt
     3781 gcgtgctgtg cttcggcttc ttgttggtgg tcgttcgctt tcaagtccac gactcctgcc
     3841 gttgctcacc gccgggctgt cttctttcgc ccctccctcg gtgcctcttc cgccgccgca
     3901 cgcgtgcgct gatcacgcgt cttgcgtgat gtgcctgtgt gtatacacac cgtgcacgga
     3961 gagagcgagc gagcgagaga gagagagggc gagaggaaga atggccggtg cctcgcgtgg
     4021 gcaagcgtgc gcgagtttgt cgtgtcgctg cgccgcgtgg atgtcgcgca gcgcaagacc
     4081 ccgtgcgcct gaaacgtgaa gggaggtcga gaagcgtgcc gcatgaggcc ttaaggcagg
     4141 aaagtgaaaa gagctcgacg cgagtgcagc acgcgcgcat cgacaggacg gaatagacac
     4201 tcagcccctt cccctttcaa aaactgaatg gacggacatc gtaacgcgct tctctcccct
     4261 ccactccccc tgtccctgtc tccttgacgt gtgctgctct cgtgagcgac agatgaagcg
     4321 aggcgtagaa ggaaaggggg ggggagcagg cctgcacaca cacacacaca cacacacaca
     4381 cacacacaca cacacacaca cacacacgaa aagcaagttt gaggcagacg tggtttcaca
     4441 cgattcctgt gcaagatgcg tgcctgcgtg acgggtgtgg ctgtgtatgc atgcgtgaag
     4501 ggttaccgca acgatgcgca atggactccc ccgctttcca tttcgtcacc ttccgcctct
     4561 ctctctctct ctctcaccat ctacgcgtgc acatcatcaa ctgtctcttg tcggtgctca
     4621 ccaccctcaa ccacccctca ctttcaaggc ttcccgaacg cacacaaaag gcgtgaaaac
     4681 cgctcgcggt gtgttgagcc gtccaccgtg ccctccccct gtccccgggg gatccactag
     4741 acgaccttcc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc
     4801 ccgggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt
     4861 cgacccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt
     4921 cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac
     4981 cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga
     5041 gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg
     5101 gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa
     5161 gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc
     5221 cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac
     5281 cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc
     5341 cggtgcctag atgcccttgt gtgcgtgtgt gtggtggttg cagcgatgcc cggcgcgtgt
     5401 gggcacctcc ttgggtgcgc gcccgccgtg gcagctgcgc gtgcgtgcga gatgtgaggc
     5461 agaggaagag gaaggcgatg cgggcgacag cgcagcgagg tgcggcggag cgtagggggg
     5521 aaatggacga gcaggcgcgc tgtgaatcgg agctgcggca ccacccaagt cgtggtgccc
     5581 cgcgaatggc tgttctgccc gccctcgctt cacgcctccc cctcccctcg cgtgccctcg
     5641 cgtggcctcc cttgttatcc ctctctctcg cacgcacacg cacacgcgta tacgcgagcc
     5701 cgctattctg ccttcgtctg gctctttgta ttttgcttgc ttcttcagca cacttgtgtg
     5761 ctgtgcgttc agcgatatct tccactactt tgttttctcc tccccctcgg gaggtgcttc
     5821 gcttgtgctt tgacggtggt gcgtggctgc tgggttatgt gccgggcgtg cgcgcctccg
     5881 ccgcctccct gcagcttgtg ggtgcgctgc gttcgcaccg cgctcgcgtg catgcacatg
     5941 cctgcactgc gtgcgggacg ccttccgggc gcgttggccc cccgcctctg cagccacggt
     6001 ctgcttattg attgtgcttg cttcatcggc tcttctctgc gcgcgtgcgt gcgtgcgtgt
     6061 gcgtgtccgt gcgtatgcgt gaggcgcaac ggtccccaga gcaaggcatg tcgaggggaa
     6121 cactatagac gcatgtgtac gtgtacacga tgtgtatacg tatacgtgta ccgaatggtg
     6181 cgtgcgcgtg tgcagcattg ccgtgacggc atgtacgaag cgctgcagtg ggatggaccc
     6241 tgtgcgcgtg ccggagaggt agtgtcgcgt gtgggtgcgg agtgatggag gccagggggc
     6301 ttacgagcac cgtcgctttt cccccgatgg cggctggcac gcagcgcacg caccggggat
     6361 gtgtgacgtg cgtcgctgtg cgcctctccc tctcccctgt gcgcgcggcg catggatgca
     6421 ccgctgttgt gtgaggttgc ccgcacctgc gttgttgcct gtgatgacgt ccctccctct
     6481 cttgcactct ccccgtcccc acctgccctg caccgtggtc gactgctccc gacgccctgc
     6541 acagactctc gtcgccacca ccagcagcag ccctctatat acccgccact gccgcagcgt
     6601 tcgggccgtg gcctctgcgt ttcacttgct ctcccctcgc tctgttcatt gcttccttct
     6661 gttcccctcg ctgcccgcgt ccgcgactag acctaacgcc ttttcaactc acggcctcta
     6721 ggaatgaagg agggtagttc gggggagaac gtactggggc gtcagaggtg aaattcttag
     6781 accgcaccaa gacgaactac agcgaaggca ttcttcaagg ataccttcct caatcaagaa
     6841 ccaaagtgtg gagatcgaag atgattagag accattgtag tccacactgc aaacgatgac
     6901 acccatgaat tggggatctt atgggccggc ctgcggcagg gtttaccctg tgtcagcacc
     6961 gcgcccgctt ttaccaactt acgtatcttt tctattcggc ctttaccggc cacccacggg
     7021 aatatcctca gcacgttttc tgttttttca cgcgaaagct ttgaggttac agtctcaggg
     7081 gggagtacgt tcgcaagagt gaaacttaaa gaaattgacg gaatggcacc acaagacgtg
     7141 gagcgtgcgg tttaatttga ctcaacacgg ggaactttac cagatccgga caggatgagg
     7201 attgacagat tgagtgttct ttctcgattc cctgaatggt ggtgcatggc cgcttttggt
     7261 cggtggagtg atttgtttgg ttgattccgt caacggacga gatccaagct gcccagtaga
     7321 attcagaatt gcccatagga tagcaaactc atcggcgggt tttacccaac ggtgggccgc
     7381 attcggtcga attcttctct gcgggattcc tttgtaattg cacaaggtga aattttgggc
     7441 aacagcaggt ctgtgatgct cctcaatgtt ctgggcgaca cgcgcactac aatgtcagtg
     7501 agaacaagaa aaacgacttt tgtcgaacct acttgatcaa aagagtgggg aaaccccgga
     7561 atcacataga cccacttggg accgaggatt gcaattattg gtcgcgcaac gaggaatgtc
     7621 tcgtaggcgc agctcatgtt taaaccagct tggcgtaatc atggtcatag ctgtttcctg
     7681 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     7741 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     7801 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     7861 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     7921 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     7981 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     8041 gtaaaaaggc cgcgttgctg gcgtt
//
