LOCUS       sequence-258264-        6108 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression plasmid for PURE system components.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6108)
  AUTHORS   Shimizu Y, Inoue A, Tomari Y, Suzuki T, Yokogawa T, Nishikawa K, 
            Ueda T
  TITLE     Cell-free translation reconstituted with purified components.
  JOURNAL   Nat Biotechnol. 2001 Aug;19(8):751-5. doi: 10.1038/90802.
  PUBMED    11479568
REFERENCE   2  (bases 1 to 6108)
  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"; journalName: 
            "Nat Biotechnol"; date: "2001-08"; volume: "19"; issue: "8"; pages: 
            "751-5"
COMMENT     Sequence Label: pQE30-His-T7RNAP
FEATURES             Location/Qualifiers
     source          1..6108
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..24,25..30,31..47,48..54)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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)."
     protein_bind    62..78
                     /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             97..108
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             127..144
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2855..2949
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             2993..3652
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3059..3078)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      3717..3803
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(3850..3872)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3958..4098
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4013..4032
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4113..4130)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4284..4872)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4364..4383)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5043..5834,5835..5903))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5666..5685
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5904..6008)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6076..6094
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgaga
      121 ggatcgcatc accatcacca tcacggatcc atgaacacga ttaacatcgc taagaacgac
      181 ttctctgaca tcgaactggc tgctatcccg ttcaacactc tggctgacca ttacggtgag
      241 cgtttagctc gcgaacagtt ggcccttgag catgagtctt acgagatggg tgaagcacgc
      301 ttccgcaaga tgtttgagcg tcaacttaaa gctggtgagg ttgcggataa cgctgccgcc
      361 aagcctctca tcactaccct actccctaag atgattgcac gcatcaacga ctggtttgag
      421 gaagtgaaag ctaagcgcgg caagcgcccg acagccttcc agttcctgca agaaatcaag
      481 ccggaagccg tagcgtacat caccattaag accactctgg cttgcctaac cagtgctgac
      541 aatacaaccg ttcaggctgt agcaagcgca atcggtcggg ccattgagga cgaggctcgc
      601 ttcggtcgta tccgtgacct tgaagctaag cacttcaaga aaaacgttga ggaacaactc
      661 aacaagcgcg tagggcacgt ctacaagaaa gcatttatgc aagttgtcga ggctgacatg
      721 ctctctaagg gtctactcgg tggcgaggcg tggtcttcgt ggcataagga agactctatt
      781 catgtaggag tacgctgcat cgagatgctc attgagtcaa ccggaatggt tagcttacac
      841 cgccaaaatg ctggcgtagt aggtcaagac tctgagacta tcgaactcgc acctgaatac
      901 gctgaggcta tcgcaacccg tgcaggtgcg ctggctggca tctctccgat gttccaacct
      961 tgcgtagttc ctcctaagcc gtggactggc attactggtg gtggctattg ggctaacggt
     1021 cgtcgtcctc tggcgctggt gcgtactcac agtaagaaag cactgatgcg ctacgaagac
     1081 gtttacatgc ctgaggtgta caaagcgatt aacattgcgc aaaacaccgc atggaaaatc
     1141 aacaagaaag tcctagcggt cgccaacgta atcaccaagt ggaagcattg tccggtcgag
     1201 gacatccctg cgattgagcg tgaagaactc ccgatgaaac cggaagacat cgacatgaat
     1261 cctgaggctc tcaccgcgtg gaaacgtgct gccgctgctg tgtaccgcaa ggacaaggct
     1321 cgcaagtctc gccgtatcag ccttgagttc atgcttgagc aagccaataa gtttgctaac
     1381 cataaggcca tctggttccc ttacaacatg gactggcgcg gtcgtgttta cgctgtgtca
     1441 atgttcaacc cgcaaggtaa cgatatgacc aaaggactgc ttacgctggc gaaaggtaaa
     1501 ccaatcggta aggaaggtta ctactggctg aaaatccacg gtgcaaactg tgcgggtgtc
     1561 gataaggttc cgttccctga gcgcatcaag ttcattgagg aaaaccacga gaacatcatg
     1621 gcttgcgcta agtctccact ggagaacact tggtgggctg agcaagattc tccgttctgc
     1681 ttccttgcgt tctgctttga gtacgctggg gtacagcacc acggcctgag ctataactgc
     1741 tcccttccgc tggcgtttga cgggtcttgc tctggcatcc agcacttctc cgcgatgctc
     1801 cgagatgagg taggtggtcg cgcggttaac ttgcttccta gtgaaaccgt tcaggacatc
     1861 tacgggattg ttgctaagaa agtcaacgag attctacaag cagacgcaat caatgggacc
     1921 gataacgaag tagttaccgt gaccgatgag aacactggtg aaatctctga gaaagtcaag
     1981 ctgggcacta aggcactggc tggtcaatgg ctggcttacg gtgttactcg cagtgtgact
     2041 aagcgttcag tcatgacgct ggcttacggg tccaaagagt tcggcttccg tcaacaagtg
     2101 ctggaagata ccattcagcc agctattgat tccggcaagg gtctgatgtt cactcagccg
     2161 aatcaggctg ctggatacat ggctaagctg atttgggaat ctgtgagcgt gacggtggta
     2221 gctgcggttg aagcaatgaa ctggcttaag tctgctgcta agctgctggc tgctgaggtc
     2281 aaagataaga agactggaga gattcttcgc aagcgttgcg ctgtgcattg ggtaactcct
     2341 gatggtttcc ctgtgtggca ggaatacaag aagcctattc agacgcgctt gaacctgatg
     2401 ttcctcggtc agttccgctt acagcctacc attaacacca acaaagatag cgagattgat
     2461 gcacacaaac aggagtctgg tatcgctcct aactttgtac acagccaaga cggtagccac
     2521 cttcgtaaga ctgtagtgtg ggcacacgag aagtacggaa tcgaatcttt tgcactgatt
     2581 cacgactcct tcggtaccat tccggctgac gctgcgaacc tgttcaaagc agtgcgcgaa
     2641 actatggttg acacatatga gtcttgtgat gtactggctg atttctacga ccagttcgct
     2701 gaccagttgc acgagtctca attggacaaa atgccagcac ttccggctaa aggtaacttg
     2761 aacctccgtg acatcttaga gtcggacttc gcgttcgcgt aactgcaggc atcgcgttcg
     2821 cgtaactgca gccaagctta attagctgag cttggactcc tgttgataga tccagtaatg
     2881 acctcagaac tccatctgga tttgttcaga acgctcggtt gccgccgggc gttttttatt
     2941 ggtgagaatc caagctagct tggcgagatt ttcaggagct aaggaagcta aaatggagaa
     3001 aaaaatcact ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga
     3061 ggcatttcag tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc
     3121 ctttttaaag accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct
     3181 tgcccgcctg atgaatgctc atccggaatt tcgtatggca atgaaagacg gtgagctggt
     3241 gatatgggat agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc
     3301 atcgctctgg agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga
     3361 tgtggcgtgt tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt
     3421 tttcgtctca gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat
     3481 ggacaacttc ttcgcccccg ttttcaccat gggcaaatat tatacgcaag gcgacaaggt
     3541 gctgatgccg ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag
     3601 aatgcttaat gaattacaac agtactgcga tgagtggcag ggcggggcgt aattttttta
     3661 aggcagttat tggtgccctt aaacgcctgg ggtaatgact ctctagcttg aggcatcaaa
     3721 taaaacgaaa ggctcagtcg aaagactggg cctttcgttt tatctgttgt ttgtcggtga
     3781 acgctctcct gagtaggaca aatccgccct ctagagctgc ctcgcgcgtt tcggtgatga
     3841 cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga
     3901 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc
     3961 agccatgacc cagtcacgta gcgatagcgg agtgtatact ggcttaacta tgcggcatca
     4021 gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg
     4081 agaaaatacc gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     4141 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     4201 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     4261 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     4321 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     4381 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     4441 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     4501 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     4561 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     4621 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     4681 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     4741 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     4801 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     4861 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     4921 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     4981 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     5041 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     5101 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     5161 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     5221 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     5281 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     5341 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     5401 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     5461 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     5521 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     5581 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     5641 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     5701 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     5761 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     5821 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     5881 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     5941 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     6001 gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg
     6061 acattaacct ataaaaatag gcgtatcacg aggccctttc gtcttcac
//
