LOCUS       sequence-176689-        6017 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  [Edit] Expresses recombinant 6HIS tagged T. cruzi cytoplasmic 
            tryparedoxin peroxidase protein in E. coli upon IPTG induction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6017)
  AUTHORS   Dotiwala F, Mulik S, Polidoro RB, Ansara JA, Burleigh BA, Walch M, 
            Gazzinelli RT, Lieberman J
  TITLE     Killer lymphocytes use granulysin, perforin and granzymes to kill 
            intracellular parasites.
  JOURNAL   Nat Med. 2016 Feb;22(2):210-6. doi: 10.1038/nm.4023. Epub 2016 Jan 
            11.
  PUBMED    26752517
REFERENCE   2  (bases 1 to 6017)
  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: "10.1038/nm.4023"; 
            journalName: "Nat Med"; date: "2016-02"; volume: "22"; issue: "2"; 
            pages: "210-6"
COMMENT     Sequence Label: pET21a - CPX
FEATURES             Location/Qualifiers
     source          1..6017
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..33
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             661..678
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(731..749)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      745..792
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      829..1284
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(916..935)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1126..1147
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1310..1414
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1415..1483,1484..2275)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1633..1652)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2446..3034
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2935..2954
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3188..3205
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3220..3359
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3285..3304)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3445..3467
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3461..3652)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4461..5543)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5505..5524
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5544..5621)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5827..5846
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5930..5949
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5930..5948
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5949..5973
                     /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)."
     RBS             5988..6010
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 atggctagca tgactggtgg acagcaaatg ggtcgcggat ccgaattcat gtcctgcgga
       61 gacgcaaagc tcaaccaccc ggcgcccgac ttcaatgaga cggcgctgat gcccaacggc
      121 accttcaaga aggtggctct cacctcctac aagggcaagt ggctggtgct cttcttctac
      181 ccgatggact tcaccttcgt ctgccccaca gagatctgcc aattctcgga ccgtgtgaag
      241 gagttctctg acattggctg cgaggtgctt gcctgctcca tggacagcga gtactcccat
      301 ctggcctgga caagcattga gcgcaagcgt ggcggacttg gccagatgaa catccccatt
      361 cttgccgaca agaccaagtg catcatgaag tcgtatggtg tgctgaagga ggaggatggc
      421 gtggcgtacc gtggcctttt catcatcgac ccgaagcaga acctgcggca gatcaccgtc
      481 aacgacctcc ccgttggccg cgacgtggac gaggcccttc gccttgtgaa ggcgttccag
      541 ttcgtggaga agcatggcga ggtgtgcccc gccaactgga agcccggtga caagacgatg
      601 aagccggatc ccgaaaagtc caaggagtac tttggtgctg tcgcggcggc cgcactcgag
      661 caccaccacc accaccactg agatccggct gctaacaaag cccgaaagga agctgagttg
      721 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg
      781 aggggttttt tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta
      841 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
      901 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
      961 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     1021 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     1081 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     1141 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     1201 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     1261 acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     1321 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     1381 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     1441 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     1501 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     1561 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     1621 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     1681 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     1741 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     1801 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     1861 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     1921 agccatacca aacgacgagc gtgacaccac gatgcctgca gcaatggcaa caacgttgcg
     1981 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     2041 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     2101 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     2161 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     2221 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     2281 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     2341 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     2401 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     2461 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     2521 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     2581 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     2641 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     2701 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     2761 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     2821 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     2881 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     2941 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     3001 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     3061 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     3121 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     3181 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct
     3241 tacgcatctg tgcggtattt cacaccgcaa tggtgcactc tcagtacaat ctgctctgat
     3301 gccgcatagt taagccagta tacactccgc tatcgctacg tgactgggtc atggctgcgc
     3361 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     3421 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     3481 caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac
     3541 agatgtctgc ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct
     3601 ggcttctgat aaagcgggcc atgttaaggg cggttttttc ctgtttggtc actgatgcct
     3661 ccgtgtaagg gggatttctg ttcatggggg taatgatacc gatgaaacga gagaggatgc
     3721 tcacgatacg ggttactgat gatgaacatg cccggttact ggaacgttgt gagggtaaac
     3781 aactggcggt atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct
     3841 tcgttaatac agatgtaggt gttccacagg gtagccagca gcatcctgcg atgcagatcc
     3901 ggaacataat ggtgcagggc gctgacttcc gcgtttccag actttacgaa acacggaaac
     3961 cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg
     4021 ttcgctcgcg tatcggtgat tcattctgct aaccagtaag gcaaccccgc cagcctagcc
     4081 gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg ggccgccatg ccggcgataa
     4141 tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg
     4201 cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc
     4261 ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg tcctacgagt tgcatgataa
     4321 agaagacagt cataagtgcg gcgacgatag tcatgccccg cgcccaccgg aaggagctga
     4381 ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg tgcctaatga gtgagctaac
     4441 ttacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     4501 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgccagggtg
     4561 gtttttcttt tcaccagtga gacgggcaac agctgattgc ccttcaccgc ctggccctga
     4621 gagagttgca gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg
     4681 gtggttaacg gcgggatata acatgagctg tcttcggtat cgtcgtatcc cactaccgag
     4741 atatccgcac caacgcgcag cccggactcg gtaatggcgc gcattgcgcc cagcgccatc
     4801 tgatcgttgg caaccagcat cgcagtggga acgatgccct cattcagcat ttgcatggtt
     4861 tgttgaaaac cggacatggc actccagtcg ccttcccgtt ccgctatcgg ctgaatttga
     4921 ttgcgagtga gatatttatg ccagccagcc agacgcagac gcgccgagac agaacttaat
     4981 gggcccgcta acagcgcgat ttgctggtga cccaatgcga ccagatgctc cacgcccagt
     5041 cgcgtaccgt cttcatggga gaaaataata ctgttgatgg gtgtctggtc agagacatca
     5101 agaaataacg ccggaacatt agtgcaggca gcttccacag caatggcatc ctggtcatcc
     5161 agcggatagt taatgatcag cccactgacg cgttgcgcga gaagattgtg caccgccgct
     5221 ttacaggctt cgacgccgct tcgttctacc atcgacacca ccacgctggc acccagttga
     5281 tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc cagactggag
     5341 gtggcaacgc caatcagcaa cgactgtttg cccgccagtt gttgtgccac gcggttggga
     5401 atgtaattca gctccgccat cgccgcttcc actttttccc gcgttttcgc agaaacgtgg
     5461 ctggcctggt tcaccacgcg ggaaacggtc tgataagaga caccggcata ctctgcgaca
     5521 tcgtataacg ttactggttt cacattcacc accctgaatt gactctcttc cgggcgctat
     5581 catgccatac cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc gacgctctcc
     5641 cttatgcgac tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc
     5701 cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg ccacggggcc
     5761 tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag cccgatcttc
     5821 cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc
     5881 ggccacgatg cgtccggcgt agaggatcga gatctcgatc ccgcgaaatt aatacgactc
     5941 actatagggg aattgtgagc ggataacaat tcccctctag aaataatttt gtttaacttt
     6001 aagaaggaga tatacat
//
