LOCUS       sequence-292609-        5951 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Mtb ATR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5951)
  AUTHORS   Ruetz M, Campanello GC, Purchal M, Shen H, McDevitt L, Gouda H, 
            Wakabayashi S, Zhu J, Rubin EJ, Warncke K, Mootha VK, Koutmos M, 
            Banerjee R
  TITLE     Itaconyl-CoA forms a stable biradical in methylmalonyl-CoA mutase 
            and derails its activity and repair.
  JOURNAL   Science. 2019 Nov 1;366(6465):589-593. doi: 10.1126/science.aay0934.
  PUBMED    31672889
REFERENCE   2  (bases 1 to 5951)
  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: "10.1126/science"; 
            journalName: "Science"; date: "2019-11-1- 1"; volume: "366"; issue: 
            "6465"; pages: "589-593"
COMMENT     Sequence Label: pMtb ATR
FEATURES             Location/Qualifiers
     source          1..5951
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             100..117
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             127..144
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(886..904)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      900..947
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      984..1439
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1071..1090)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1281..1302
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1531..2346)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1531..2346)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2254..2273
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2338..2357
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2468..3056
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2957..2976
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3210..3227
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3242..3381
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3307..3326)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3467..3489
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3483..3674)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4483..5565)
                     /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     5527..5546
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5566..5643)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5849..5868
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg ggcagcagcc atcatcatca tcatcacagc
      121 agcggcctgg tgccgcgcgg cagccatatg gcagtccacc tgacccgcat ctatacccga
      181 accggcgatg acggaacgac gggattgagc gatatgtccc gtgtcgccaa aaccgacgcc
      241 cgactggtgg catacgcgga ttgcgacgag gccaacgccg cgatcggggc cgccctggcg
      301 ctgggccacc cggacactca aatcacggat gtgctgcggc agatccagaa cgacttgttc
      361 gatgccggtg cggatctgtc gactccgata gtggaaaacc cgaagcaccc tccgctgcga
      421 atcgcacagt cctatatcga tcggctcgaa gggtggtgcg acgcatacaa cgcgggcctg
      481 cctgcactaa aatcctttgt actacctggt ggttcaccgc tgtcggcgct gttgcacgtc
      541 gctcgcaccg tcgttcgccg ggccgagcga tccgcgtggg ccgcggtcga cgctcacccg
      601 gaaggcgtca gcgtactgcc ggcgaaatac ctgaatcgtc tgtcggatct gctgttcatc
      661 ctgtcgcggg tagccaatcc cgacggtgac gtgctgtggc ggcccggtgg tgataggacg
      721 gcgagctaag ctagccagat atgcagccat gaaggagggg aatcaagaga aaatatacaa
      781 gaaaaatgac ccatcggccg agtctgaggg actctgacta ccaccaccac cactgagatc
      841 cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac
      901 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa
      961 ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     1021 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     1081 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     1141 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     1201 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     1261 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     1321 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     1381 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
     1441 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     1501 tcaaatatgt atccgctcat gaattaattc ttagaaaaac tcatcgagca tcaaatgaaa
     1561 ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa
     1621 tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc
     1681 gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt
     1741 atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagtttatg
     1801 catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc
     1861 atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata cgcgatcgct
     1921 gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc
     1981 atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg ctgttttccc
     2041 ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt
     2101 cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt
     2161 ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa
     2221 tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa
     2281 atcagcatcc atgttggaat ttaatcgcgg cctagagcaa gacgtttccc gttgaatatg
     2341 gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg ttcatgacca
     2401 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     2461 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     2521 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     2581 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     2641 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     2701 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     2761 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     2821 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     2881 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     2941 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     3001 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     3061 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     3121 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     3181 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     3241 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac
     3301 tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta
     3361 cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     3421 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     3481 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca
     3541 gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt
     3601 ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt
     3661 tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata
     3721 ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta
     3781 ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc
     3841 actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag
     3901 cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc
     3961 agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt
     4021 ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta
     4081 aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat gcgcacccgt
     4141 ggggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt ggcgggacca
     4201 gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga caggccgatc
     4261 atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga cccagagcgc tgccggcacc
     4321 tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat agtcatgccc
     4381 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     4441 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     4501 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     4561 ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt
     4621 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
     4681 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
     4741 atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact cggtaatggc
     4801 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
     4861 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
     4921 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
     4981 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
     5041 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
     5101 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
     5161 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
     5221 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
     5281 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
     5341 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
     5401 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
     5461 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
     5521 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
     5581 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc attcgatggt
     5641 gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag cccagtagta
     5701 ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag atggcgccca
     5761 acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg ctcatgagcc
     5821 cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg ccagcaaccg
     5881 cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc gagatctcga
     5941 tcccgcgaaa t
//
