LOCUS       sequence-258679-        3642 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 3642)
  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 3642)
  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-IF1
FEATURES             Location/Qualifiers
     source          1..3642
                     /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      389..483
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             527..1186
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(593..612)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      1251..1337
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(1384..1406)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    1492..1632
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     1547..1566
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1647..1664)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1818..2406)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1898..1917)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2577..3368,3369..3437))
                     /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     3200..3219
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3438..3542)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3610..3628
                     /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 atggccaaag aagacaatat tgaaatgcaa
      181 ggtaccgttc ttgaaacgtt gcctaatacc atgttccgcg tagagttaga aaacggtcac
      241 gtggttactg cacacatctc cggtaaaatg cgcaaaaact acatccgcat cctgacgggc
      301 gacaaagtga ctgttgaact gaccccgtac gacctgagca aaggccgcat tgtcttccgt
      361 agtcgctgaa gctaattagc tgagcttgga ctcctgttga tagatccagt aatgacctca
      421 gaactccatc tggatttgtt cagaacgctc ggttgccgcc gggcgttttt tattggtgag
      481 aatccaagct agcttggcga gattttcagg agctaaggaa gctaaaatgg agaaaaaaat
      541 cactggatat accaccgttg atatatccca atggcatcgt aaagaacatt ttgaggcatt
      601 tcagtcagtt gctcaatgta cctataacca gaccgttcag ctggatatta cggccttttt
      661 aaagaccgta aagaaaaata agcacaagtt ttatccggcc tttattcaca ttcttgcccg
      721 cctgatgaat gctcatccgg aatttcgtat ggcaatgaaa gacggtgagc tggtgatatg
      781 ggatagtgtt cacccttgtt acaccgtttt ccatgagcaa actgaaacgt tttcatcgct
      841 ctggagtgaa taccacgacg atttccggca gtttctacac atatattcgc aagatgtggc
      901 gtgttacggt gaaaacctgg cctatttccc taaagggttt attgagaata tgtttttcgt
      961 ctcagccaat ccctgggtga gtttcaccag ttttgattta aacgtggcca atatggacaa
     1021 cttcttcgcc cccgttttca ccatgggcaa atattatacg caaggcgaca aggtgctgat
     1081 gccgctggcg attcaggttc atcatgccgt ttgtgatggc ttccatgtcg gcagaatgct
     1141 taatgaatta caacagtact gcgatgagtg gcagggcggg gcgtaatttt tttaaggcag
     1201 ttattggtgc ccttaaacgc ctggggtaat gactctctag cttgaggcat caaataaaac
     1261 gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc
     1321 tcctgagtag gacaaatccg ccctctagag ctgcctcgcg cgtttcggtg atgacggtga
     1381 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     1441 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat
     1501 gacccagtca cgtagcgata gcggagtgta tactggctta actatgcggc atcagagcag
     1561 attgtactga gagtgcacca tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa
     1621 taccgcatca ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     1681 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     1741 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     1801 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     1861 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     1921 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     1981 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     2041 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     2101 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     2161 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     2221 ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct
     2281 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     2341 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     2401 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     2461 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     2521 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     2581 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     2641 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     2701 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     2761 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     2821 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     2881 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     2941 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     3001 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     3061 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     3121 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     3181 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     3241 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     3301 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     3361 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     3421 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     3481 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     3541 cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta
     3601 acctataaaa ataggcgtat cacgaggccc tttcgtcttc ac
//
