LOCUS       sequence-176709-        5999 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses recombinant 6HIS tagged T. cruzi superoxide dismutase B 
            protein in E. coli upon IPTG induction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5999)
  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 5999)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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 - SodB
FEATURES             Location/Qualifiers
     source          1..5999
                     /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             643..660
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(713..731)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      727..774
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      811..1266
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(898..917)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1108..1129
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1292..1396
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1397..1465,1466..2257)
                     /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(1615..1634)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2428..3016
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2917..2936
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3170..3187
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3202..3341
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3267..3286)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3427..3449
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3443..3634)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4443..5525)
                     /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     5487..5506
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5526..5603)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5809..5828
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5912..5931
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5912..5930
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5931..5955
                     /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             5970..5992
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 atggctagca tgactggtgg acagcaaatg ggtcgcggat ccatggtctt cagcattcct
       61 ccgctcccat ggggctacga tgggcttgcg gcaaaaggcc tctcaaagca gcaggtgacg
      121 ctccactacg acaagcacca tcaggggtat gtgacgaaac tcaacgccgc ggcgcagaca
      181 aactccgcgc ttgcaacgaa gagcatcgag gagatcatca ggacggagaa aggccccatc
      241 ttcaaccttg cggcgcagat ttttaaccac acgttctact gggagagcat gtgtcctaat
      301 ggcggtggcg agccgacggg aaaagttgcc gacgagatca acgcttcatt tggcagtttt
      361 gcgaagttca aggaggagtt tacaaacgtg gctgtgggcc actttggctc gggttgggcg
      421 tggcttgtga aggacaccaa ttccggcaaa ctgaaagtct accagacgca tgacgcggga
      481 tgtccactga cagagcccaa cttgaagcct ctccttacat gcgatgtatg ggagcatgcg
      541 tactacgtgg actacaagaa cgaccgtgcg gcatatgttc agaccttttg gaacgttgtc
      601 aactggaaga acgtggaacg gcaacttgcg gccgcactcg agcaccacca ccaccaccac
      661 tgagatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag
      721 caataactag cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa
      781 ggaggaacta tatccggatt ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg
      841 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
      901 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
      961 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     1021 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     1081 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     1141 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     1201 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     1261 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     1321 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     1381 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     1441 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     1501 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     1561 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     1621 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     1681 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     1741 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     1801 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     1861 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     1921 gcgtgacacc acgatgcctg cagcaatggc aacaacgttg cgcaaactat taactggcga
     1981 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     2041 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     2101 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     2161 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     2221 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     2281 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     2341 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     2401 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     2461 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     2521 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
     2581 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     2641 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     2701 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     2761 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     2821 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     2881 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     2941 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     3001 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     3061 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     3121 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     3181 gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     3241 ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     3301 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
     3361 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     3421 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
     3481 agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat
     3541 ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg
     3601 ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc
     3661 tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg
     3721 atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc
     3781 ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag
     3841 gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg
     3901 gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg
     3961 ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg
     4021 attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga
     4081 gcacgatcat gcgcacccgt ggggccgcca tgccggcgat aatggcctgc ttctcgccga
     4141 aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata
     4201 ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga
     4261 cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg
     4321 cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca
     4381 agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc
     4441 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     4501 aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt
     4561 gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg
     4621 tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata
     4681 taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc
     4741 agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc
     4801 atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg
     4861 gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta
     4921 tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg
     4981 atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg
     5041 gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca
     5101 ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc
     5161 agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg
     5221 cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc
     5281 gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc
     5341 aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc
     5401 atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg
     5461 cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt
     5521 ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag
     5581 gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat
     5641 taggaagcag cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc
     5701 atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc
     5761 gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc
     5821 gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc
     5881 gtagaggatc gagatctcga tcccgcgaaa ttaatacgac tcactatagg ggaattgtga
     5941 gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga gatatacat
//
