LOCUS       sequence-176706-        5846 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  [Edit] Expresses recombinant 6HIS tagged T. cruzi tryparedoxin 
            protein in E. coli upon IPTG induction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5846)
  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 5846)
  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 - TryX
FEATURES             Location/Qualifiers
     source          1..5846
                     /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             490..507
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(560..578)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      574..621
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      658..1113
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(745..764)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     955..976
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1139..1243
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1244..1312,1313..2104)
                     /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(1462..1481)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2275..2863
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2764..2783
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3017..3034
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3049..3188
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3114..3133)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3274..3296
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3290..3481)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4290..5372)
                     /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     5334..5353
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5373..5450)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5656..5675
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5759..5778
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5759..5777
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5778..5802
                     /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             5817..5839
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 atggctagca tgactggtgg acagcaaatg ggtcgcggat ccatgtctgg tttggcgaag
       61 tacctcccca gcaccattaa tctggtctcc aagagcggca ccgtcagccc catttccctt
      121 gctgggaaga cggtgttttt ttacttctcc gcctcctggt gcccgccctg ccgcggcttc
      181 accccgaccc ttgtggagtt ttacgagaag ttccgtgaat cgaagaactt cgaggtcgta
      241 cttgtcacct gggacgacga ggaggaggcg tacaacgggt acttcgccaa gatgccatgg
      301 ctcgccatcc ccttctcgag ccgcacagag ctggaggccc tccggtcgac ttttggcgtg
      361 gagactatcc ccacagttat cgccgtcaac gccgacacgg gcgccgtggt cagcacgaag
      421 ggacgcgagc gacttttgac cgaccccgag ggcaaaaact tcccctggtc cgacgcggcc
      481 gcactcgagc accaccacca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa
      541 gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
      601 cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc gaatgggacg
      661 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
      721 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
      781 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
      841 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
      901 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
      961 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     1021 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     1081 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     1141 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     1201 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     1261 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     1321 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     1381 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     1441 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     1501 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     1561 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     1621 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     1681 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     1741 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag caatggcaac
     1801 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     1861 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     1921 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     1981 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     2041 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     2101 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     2161 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     2221 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     2281 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     2341 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     2401 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     2461 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     2521 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     2581 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     2641 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     2701 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     2761 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     2821 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     2881 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     2941 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     3001 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     3061 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
     3121 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     3181 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     3241 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     3301 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     3361 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     3421 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     3481 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     3541 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     3601 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     3661 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     3721 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     3781 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     3841 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     3901 agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc
     3961 cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt
     4021 gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc
     4081 agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt
     4141 gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga
     4201 aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag
     4261 tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4321 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4381 gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc
     4441 tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc
     4501 tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc
     4561 actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc
     4621 agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt
     4681 tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc
     4741 tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca
     4801 gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc
     4861 acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca
     4921 gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc
     4981 tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc
     5041 accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca
     5101 cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc
     5161 agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg
     5221 cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca
     5281 gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac
     5341 tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg actctcttcc
     5401 gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg
     5461 acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt
     5521 gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc
     5581 cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc
     5641 ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc
     5701 ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc cgcgaaatta
     5761 atacgactca ctatagggga attgtgagcg gataacaatt cccctctaga aataattttg
     5821 tttaacttta agaaggagat atacat
//
