LOCUS       sequence-370660-        6144 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses secreted Mycobacterium tuberculosis (M. tb) chorismate 
            mutase (CM) containing a C-terminal biotinylation site and His tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6144)
  AUTHORS   van Neer RHP, Dranchak PK, Aitha M, Liu L, Carlson EK, Jacobsen IE, 
            Battaile K, Fang Y, Tao D, Rai G, Padia J, Lovell S, Suga H, Inglese
            J
  TITLE     Active- and Allosteric-Site Cyclic Peptide Inhibitors of Secreted M.
            tuberculosis Chorismate Mutase.
  JOURNAL   ACS Infect Dis. 2025 Mar 14;11(3):703-714. doi: 
            10.1021/acsinfecdis.4c00798. Epub 2025 Feb 4.
  PUBMED    39903128
REFERENCE   2  (bases 1 to 6144)
  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: 
            "10.1021/acsinfecdis.4c00798"; journalName: "ACS Infect Dis"; date: 
            "2025-03-14- 14"; volume: "11"; issue: "3"; pages: "703-714"
COMMENT     Sequence Label: *MtbCM-bio-His
FEATURES             Location/Qualifiers
     source          1..6144
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(191..220)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             complement(230..247)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(257..301)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             complement(908..940)
                     /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"
     RBS             948..970
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    985..1009
                     /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)."
     primer_bind     complement(1009..1028)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1010..1028)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1112..1131)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1337..1414
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1415..2497
                     /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"
     CDS             3306..3497
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3491..3513)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3599..3741
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3654..3673
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3756..3773)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3927..4515)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4007..4026)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4686..5477,5478..5546))
                     /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     5309..5328
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5547..5651)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5678..6133)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5815..5836)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6027..6046
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagctttt
      181 attagagctc atggtggtgg tggtgatggt ggtggtggtg acccgccgca gaaccacgcg
      241 gaaccagacc agaggattcg tgccattcga ttttctgagc ttcgaagatg tcgttcagac
      301 ccgccggcgg cagagcctgg caataagact gggtcgcggt ggtcagcgca cgctggtaca
      361 gagagtccag gtggcgagaa cgaacgatgt cgcgttttgc acggtccagc tgagccgcgc
      421 aagacggcgc agacagcaga gaccagtgag accagatctg agacagcata cggttgttca
      481 gagagtcgat cgcggagcgg gacgcgctca ggtccggcgg ttccggcggt gcagacgccg
      541 ggttcagttt ccagtcagag aaacgagagt attcgatcgc ttcggtagca cggatctggt
      601 cgtcgaaaac acgggtaacg tagtccgggt cgatgtgctg agaacgcgca tcttcaccca
      661 gtttcgccag ctgctgttca acacgaccag aatcttcgat cggcagctgc gcacgccatt
      721 tgaacgcagc caccgggtcc gcaacttcca gacgttccgc agccgcgtca accagttccg
      781 ccagctgaga ggtgccgtcc gcacgcgcca gcggcggacc cagcggagcg atcagagaca
      841 gcaggatacc gatagaaacc gcggtagcca ggtagatttc acgcggacgg gtcagcatgg
      901 atccgcgacc catttgctgt ccaccagtca tgctagccat atgtatatct ccttcttaaa
      961 gttaaacaaa attatttcta gaggggaatt gttatccgct cacaattccc ctatagtgag
     1021 tcgtattaat ttcgcgggat cgagatctcg atcctctacg ccggacgcat cgtggccggc
     1081 atcaccggcg ccacaggtgc ggttgctggc gcctatatcg ccgacatcac cgatggggaa
     1141 gatcgggctc gccacttcgg gctcatgagc gcttgtttcg gcgtgggtat ggtggcaggc
     1201 cccgtggccg ggggactgtt gggcgccatc tccttgcatg caccattcct tgcggcggcg
     1261 gtgctcaacg gcctcaacct actactgggc tgcttcctaa tgcaggagtc gcataaggga
     1321 gagcgtcgag atcccggaca ccatcgaatg gcgcaaaacc tttcgcggta tggcatgata
     1381 gcgcccggaa gagagtcaat tcagggtggt gaatgtgaaa ccagtaacgt tatacgatgt
     1441 cgcagagtat gccggtgtct cttatcagac cgtttcccgc gtggtgaacc aggccagcca
     1501 cgtttctgcg aaaacgcggg aaaaagtgga agcggcgatg gcggagctga attacattcc
     1561 caaccgcgtg gcacaacaac tggcgggcaa acagtcgttg ctgattggcg ttgccacctc
     1621 cagtctggcc ctgcacgcgc cgtcgcaaat tgtcgcggcg attaaatctc gcgccgatca
     1681 actgggtgcc agcgtggtgg tgtcgatggt agaacgaagc ggcgtcgaag cctgtaaagc
     1741 ggcggtgcac aatcttctcg cgcaacgcgt cagtgggctg atcattaact atccgctgga
     1801 tgaccaggat gccattgctg tggaagctgc ctgcactaat gttccggcgt tatttcttga
     1861 tgtctctgac cagacaccca tcaacagtat tattttctcc catgaagacg gtacgcgact
     1921 gggcgtggag catctggtcg cattgggtca ccagcaaatc gcgctgttag cgggcccatt
     1981 aagttctgtc tcggcgcgtc tgcgtctggc tggctggcat aaatatctca ctcgcaatca
     2041 aattcagccg atagcggaac gggaaggcga ctggagtgcc atgtccggtt ttcaacaaac
     2101 catgcaaatg ctgaatgagg gcatcgttcc cactgcgatg ctggttgcca acgatcagat
     2161 ggcgctgggc gcaatgcgcg ccattaccga gtccgggctg cgcgttggtg cggatatctc
     2221 ggtagtggga tacgacgata ccgaagacag ctcatgttat atcccgccgt taaccaccat
     2281 caaacaggat tttcgcctgc tggggcaaac cagcgtggac cgcttgctgc aactctctca
     2341 gggccaggcg gtgaagggca atcagctgtt gcccgtctca ctggtgaaaa gaaaaaccac
     2401 cctggcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct
     2461 ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtaagtt
     2521 agctcactca ttaggcaccg ggatctcgac cgatgccctt gagagccttc aacccagtca
     2581 gctccttccg gtgggcgcgg ggcatgacta tcgtcgccgc acttatgact gtcttcttta
     2641 tcatgcaact cgtaggacag gtgccggcag cgctctgggt cattttcggc gaggaccgct
     2701 ttcgctggag cgcgacgatg atcggcctgt cgcttgcggt attcggaatc ttgcacgccc
     2761 tcgctcaagc cttcgtcact ggtcccgcca ccaaacgttt cggcgagaag caggccatta
     2821 tcgccggcat ggcggcccca cgggtgcgca tgatcgtgct cctgtcgttg aggacccggc
     2881 taggctggcg gggttgcctt actggttagc agaatgaatc accgatacgc gagcgaacgt
     2941 gaagcgactg ctgctgcaaa acgtctgcga cctgagcaac aacatgaatg gtcttcggtt
     3001 tccgtgtttc gtaaagtctg gaaacgcgga agtcagcgcc ctgcaccatt atgttccgga
     3061 tctgcatcgc aggatgctgc tggctaccct gtggaacacc tacatctgta ttaacgaagc
     3121 gctggcattg accctgagtg atttttctct ggtcccgccg catccatacc gccagttgtt
     3181 taccctcaca acgttccagt aaccgggcat gttcatcatc agtaacccgt atcgtgagca
     3241 tcctctctcg tttcatcggt atcattaccc ccatgaacag aaatccccct tacacggagg
     3301 catcagtgac caaacaggaa aaaaccgccc ttaacatggc ccgctttatc agaagccaga
     3361 cattaacgct tctggagaaa ctcaacgagc tggacgcgga tgaacaggca gacatctgtg
     3421 aatcgcttca cgaccacgct gatgagcttt accgcagctg cctcgcgcgt ttcggtgatg
     3481 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
     3541 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg
     3601 cagccatgac ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc
     3661 agagcagatt gtactgagag tgcaccatat atgcggtgtg aaataccgca cagatgcgta
     3721 aggagaaaat accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     3781 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     3841 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     3901 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     3961 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     4021 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     4081 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     4141 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     4201 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     4261 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     4321 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt
     4381 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     4441 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     4501 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     4561 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     4621 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     4681 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     4741 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     4801 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     4861 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     4921 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     4981 cgcaacgttg ttgccattgc tgcaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     5041 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     5101 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     5161 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     5221 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     5281 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     5341 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     5401 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     5461 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     5521 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     5581 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     5641 ggggttccgc gcacatttcc ccgaaaagtg ccacctgaaa ttgtaaacgt taatattttg
     5701 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     5761 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
     5821 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
     5881 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
     5941 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     6001 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     6061 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     6121 ctacagggcg cgtcccattc gcca
//
