LOCUS       sequence-199059-        7966 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of PknG in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7966)
  AUTHORS   Walburger A, Koul A, Ferrari G, Nguyen L, Prescianotto-Baschong C, 
            Huygen K, Klebl B, Thompson C, Bacher G, Pieters J
  TITLE     Protein kinase G from pathogenic mycobacteria promotes survival 
            within macrophages.
  JOURNAL   Science. 2004 Jun 18;304(5678):1800-4. Epub 2004 May 20.
  PUBMED    15155913
REFERENCE   2  (bases 1 to 7966)
  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"; journalName: "Science."; 
            date: "2004-06-18"; volume: "304(5678)"; pages: "1800-4. Epub 2004 
            May 20"
COMMENT     Sequence Label: pET15b-pknG
FEATURES             Location/Qualifiers
     source          1..7966
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(984..1001)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(1011..1028)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1047..1069
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1084..1108
                     /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(1108..1127)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1109..1127)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1211..1230)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1436..1513
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     complement(1533..1552)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3907..4098
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4092..4114)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4200..4339
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4255..4274
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4354..4371)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4525..5113)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4605..4624)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5284..6075,6076..6144))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5907..5926
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6145..6249)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6317..6335
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(6362..6367,6368..6384,6385..6390)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     terminator      complement(6565..6612)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     6608..6626
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
ORIGIN
        1 tcggtggcca ccgtcggacc ggtccgcacg atctcgggcg cctggaagcc tggggtcccg
       61 tagaggtagc cgaacgagtt gatccgcgat accgcgccca ggtcgatcag cttgagctgt
      121 tcctcggtca gcatgatgtt ttccggcttc aggtcgttgt agaccaagcc gatggaatgc
      181 aggtagctca gcgccggcag gatctccagc aggtaggcga tggcctccgc gacgggcagt
      241 ttctgaccct tgctgcgttt gagcgattgc ccgccgacgt attccatcac gatgtagccg
      301 accggatccc cgtgcctgtc ggtgtgctcg acaaagttga agatctgcac gatcgacggg
      361 tgcaccacct cggccaggaa ctggcgttcg gccatcgcca ttgcctgcgc ttcggcatca
      421 ccggaatgca ccaggccctt gagcaccacc ggacggccgt tgacattgcg gtcgagagcg
      481 aggtagatcc agcccagtcc gccgtgcgcg atgcagcctt tgacctcgta ctggccggcg
      541 acgatgtccc cgggatttag ctgcggcagg aacgaatacg ggctgccgca atagggacac
      601 cagccctctg aagctccctt ggtctccgag tcggaccggc cgacgggacg tccacagttc
      661 cagcagaacc gcttggactc cggcaccacc gggttggtca tcagggcctc aagcggatcg
      721 atatcgggcg cccgcgggat ttccaccagg ccgccgccca gccgtctgac cggtgggcgc
      781 acccggctgg tggtggccat ccggtcttgc ggctcggtgt ccgggccgag cgtcggatgg
      841 gggaagttgt cctcatcgcc gaaatcgggg cggaacaccg cctgggtgct caggggtcga
      901 accgtcgcgg acgtcgcggt ctgggcgtcc gccggttggg tgccggggcc cgaacgttcg
      961 gtctctgacg ctttggccat atggctgccg cgcggcacca ggccgctgct gtgatgatga
     1021 tgatgatggc tgctgcccat ggtatatctc cttcttaaag ttaaacaaaa ttatttctag
     1081 aggggaattg ttatccgctc acaattcccc tatagtgagt cgtattaatt tcgcgggatc
     1141 gagatctcga tcctctacgc cggacgcatc gtggccggca tcaccggcgc cacaggtgcg
     1201 gttgctggcg cctatatcgc cgacatcacc gatggggaag atcgggctcg ccacttcggg
     1261 ctcatgagcg cttgtttcgg cgtgggtatg gtggcaggcc ccgtggccgg gggactgttg
     1321 ggcgccatct ccttgcatgc accattcctt gcggcggcgg tgctcaacgg cctcaaccta
     1381 ctactgggct gcttcctaat gcaggagtcg cataagggag agcgtcgaga tcccggacac
     1441 catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag cgcccggaag agagtcaatt
     1501 cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc gcagagtatg ccggtgtctc
     1561 ttatcagacc gtttcccgcg tggtgaacca ggccagccac gtttctgcga aaacgcggga
     1621 aaaagtggaa gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg cacaacaact
     1681 ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc agtctggccc tgcacgcgcc
     1741 gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca gcgtggtggt
     1801 gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca atcttctcgc
     1861 gcaacgcgtc agtgggctga tcattaacta tccgctggat gaccaggatg ccattgctgt
     1921 ggaagctgcc tgcactaatg ttccggcgtt atttcttgat gtctctgacc agacacccat
     1981 caacagtatt attttctccc atgaagacgg tacgcgactg ggcgtggagc atctggtcgc
     2041 attgggtcac cagcaaatcg cgctgttagc gggcccatta agttctgtct cggcgcgtct
     2101 gcgtctggct ggctggcata aatatctcac tcgcaatcaa attcagccga tagcggaacg
     2161 ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg
     2221 catcgttccc actgcgatgc tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc
     2281 cattaccgag tccgggctgc gcgttggtgc ggatatctcg gtagtgggat acgacgatac
     2341 cgaagacagc tcatgttata tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct
     2401 ggggcaaacc agcgtggacc gcttgctgca actctctcag ggccaggcgg tgaagggcaa
     2461 tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac
     2521 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagt ttcccgactg
     2581 gaaagcgggc agtgagcgca acgcaattaa tgtaagttag ctcactcatt aggcaccggg
     2641 atctcgaccg atgcccttga gagccttcaa cccagtcagc tccttccggt gggcgcgggg
     2701 catgactatc gtcgccgcac ttatgactgt cttctttatc atgcaactcg taggacaggt
     2761 gccggcagcg ctctgggtca ttttcggcga ggaccgcttt cgctggagcg cgacgatgat
     2821 cggcctgtcg cttgcggtat tcggaatctt gcacgccctc gctcaagcct tcgtcactgg
     2881 tcccgccacc aaacgtttcg gcgagaagca ggccattatc gccggcatgg cggccgacgc
     2941 gctgggctac gtcttgctgg cgttcgcgac gcgaggctgg atggccttcc ccattatgat
     3001 tcttctcgct tccggcggca tcgggatgcc cgcgttgcag gccatgctgt ccaggcaggt
     3061 agatgacgac catcagggac agcttcaagg atcgctcgcg gctcttacca gcctaacttc
     3121 gatcattgga ccgctgatcg tcacggcgat ttatgccgcc tcggcgagca catggaacgg
     3181 gttggcatgg attgtaggcg ccgccctata ccttgtctgc ctccccgcgt tgcgtcgcgg
     3241 tgcatggagc cgggccacct cgacctgaat ggaagccggc ggcacctcgc taacggattc
     3301 accactccaa gaattggagc caatcaattc ttgcggagaa ctgtgaatgc gcaaaccaac
     3361 ccttggcaga acatatccat cgcgtccgcc atctccagca gccgcacgcg gcgcatctcg
     3421 ggcagcgttg ggtcctggcc acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg
     3481 ctaggctggc ggggttgcct tactggttag cagaatgaat caccgatacg cgagcgaacg
     3541 tgaagcgact gctgctgcaa aacgtctgcg acctgagcaa caacatgaat ggtcttcggt
     3601 ttccgtgttt cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg
     3661 atctgcatcg caggatgctg ctggctaccc tgtggaacac ctacatctgt attaacgaag
     3721 cgctggcatt gaccctgagt gatttttctc tggtcccgcc gcatccatac cgccagttgt
     3781 ttaccctcac aacgttccag taaccgggca tgttcatcat cagtaacccg tatcgtgagc
     3841 atcctctctc gtttcatcgg tatcattacc cccatgaaca gaaatccccc ttacacggag
     3901 gcatcagtga ccaaacagga aaaaaccgcc cttaacatgg cccgctttat cagaagccag
     3961 acattaacgc ttctggagaa actcaacgag ctggacgcgg atgaacaggc agacatctgt
     4021 gaatcgcttc acgaccacgc tgatgagctt taccgcagct gcctcgcgcg tttcggtgat
     4081 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     4141 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     4201 gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat
     4261 cagagcagat tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca gatgcgtaag
     4321 gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4381 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4441 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4501 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4561 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4621 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4681 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4741 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4801 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4861 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4921 tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat
     4981 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     5041 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     5101 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     5161 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5221 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5281 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5341 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5401 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5461 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5521 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5581 caacgttgtt gccattgctg caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5641 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5701 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5761 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5821 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     5881 ttgctcttgc ccggcgtcaa cacgggataa taccgcgcca catagcagaa ctttaaaagt
     5941 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6001 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6061 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6121 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6181 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6241 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     6301 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcaag aattctcatg
     6361 tttgacagct tatcatcgat aagctttaat gcggtagttt atcacagtta aattgctaac
     6421 gcagtcaggc accgtgtatg aaatctaaca atgcgctcat cgtcatcctc ggcaccgtca
     6481 ccctggatgc tgtaggcata ggcttggtta tgccggtact gccgggcctc ttgcgggata
     6541 tccggatata gttcctcctt tcagcaaaaa acccctcaag acccgtttag aggccccaag
     6601 gggttatgct agttattgct cagcggtggc agcagccaac tcagcttcct ttcgggcttt
     6661 gttagcagcc ggatcctcga gcatatgaag cttttagaac gtgctggtgg gccggacctt
     6721 gttggccatg tccaccagcg tgtagcgatg ccgttgagtg ggagctaccc gggccaggct
     6781 gcgcagtgac gcctcgacac ccagccgcag cccgtgactg gtgaacggga aaccgaggat
     6841 gtggttggtg ctggccttgt tgtccttcag ccagtccagc gcgccaccca gcaccagggc
     6901 gcggatctgc agcacgcgtg gttcggtcgg gggcagcgcc tccactcttc gggcggcgtc
     6961 gcggatctgt tcctcggtga cttcactcgt tgaccggccg gacaacagag tcaccgcgct
     7021 ggtcagccgt gccgtggtga aatgccgaga agtgggcggt acctcgtcga gcgtgcgcac
     7081 ggcgccgacc cgatcacctt cggccgaccg ggctctggcc agtccgaaag ccgccgagat
     7141 cacgccgtcg ttggtgctcc acaccgtctg atagaacttg tgttcgtcgg tgttgccggc
     7201 tagttcggcg gtggcggcca gggcgagctt gggcgccagc tcgccgggaa aggtatccag
     7261 cacctcggtg aaatgtttgg tggccgagtc atagtcgccg gtgagcagct cggcgacggc
     7321 ccggtaccag accaatcgcc atcgccagcc aacgcgttcg gccagatcgt cgagttttcg
     7381 ggtggccttg gccacatcgc cgagatccag cagcgcgcgg acttccatta gcggcagctc
     7441 cactgactcg gagaagtcga cgccgtcggc gtccagcgca ccgtggcggg ccgcgcgcag
     7501 cgagtctagg gtctgcaccg gctgggagag caccgtggcc tgcaggaccg aagctgcgac
     7561 gtcggtcgga tcgaccagcg gcaccgacag cgcggtcacg atctcgttgg cggtcagctt
     7621 ctccgcgtgc acctgcccgt ccagatacac gtcggtgtgc gccaccagca ggtccactcc
     7681 aaatgtcgac cgactgggac tgaagatcgt tgatagccct ggccgcggca ccccggtgtc
     7741 ctgggcgacc acctcccgca acacgcccgt caattgcgcg gacatctctt cggcggtggt
     7801 gaaccgttgc cgcggatcgg ggtcgatggc cctgcgcagc aaccggccgt aagagtcgta
     7861 ggttttcagc accgggtcgt cttcgggtag cccatccaca taacggccat tgcgggtggg
     7921 caggtccagc gtgagcgccg cgagcgtgcg tcccacggtg tagatg
//
