LOCUS       sequence-258682-        3968 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 3968)
  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 3968)
  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-IF3
FEATURES             Location/Qualifiers
     source          1..3968
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(602..1393,1394..1462))
                     /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     1225..1244
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1463..1567)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1635..1653
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(1677..1691,1692..1697,1698..1714,1715..1721)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    1697..1713
                     /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    1729..1745
                     /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             1764..1775
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1794..1811
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2310..2321
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     terminator      2382..2476
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             2520..3179
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2586..2605)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      3244..3330
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(3377..3399)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3485..3625
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3540..3559
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3640..3657)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(3811..3968,1..431))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3891..3910)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
       61 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
      121 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
      181 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
      241 cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct
      301 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
      361 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
      421 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
      481 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
      541 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
      601 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
      661 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
      721 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
      781 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
      841 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
      901 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
      961 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     1021 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     1081 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     1141 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     1201 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     1261 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     1321 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     1381 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     1441 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     1501 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     1561 ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga
     1621 cattaaccta taaaaatagg cgtatcacga ggccctttcg tcttcacctc gagaaatcat
     1681 aaaaaattta tttgctttgt gagcggataa caattataat agattcaatt gtgagcggat
     1741 aacaatttca cacagaattc attaaagagg agaaattaac tatgagagga tcgcatcacc
     1801 atcaccatca cggatccatt aaaggcggaa aacgagttca aacggcgcgc cctaaccgta
     1861 tcaatggcga aattcgcgcc caggaagttc gcttaacagg tctggaaggc gagcagcttg
     1921 gtattgtgag tctgagagaa gctctggaga aagcagaaga agccggagta gacttagtcg
     1981 agatcagccc taacgccgag ccgccggttt gtcgtataat ggattacggc aaattcctct
     2041 atgaaaagag caagtcttct aaggaacaga agaaaaagca aaaagttatc caggttaagg
     2101 aaattaaatt ccgtcctggt acagatgaag gcgactatca ggtaaaactc cgcagcctga
     2161 ttcgctttct cgaagagggt gataaagcca aaatcacgct gcgtttccgc ggtcgtgaga
     2221 tggcgcacca gcaaatcggt atggaagtgc ttaatcgcgt gaaagacgat ttgcaagaac
     2281 tggcagtggt cgaatccttc ccaacgaaga tcgaaggccg ccagatgatc atggtgctcg
     2341 ctcctaagaa gaaacagtaa aagcttaatt agctgagctt ggactcctgt tgatagatcc
     2401 agtaatgacc tcagaactcc atctggattt gttcagaacg ctcggttgcc gccgggcgtt
     2461 ttttattggt gagaatccaa gctagcttgg cgagattttc aggagctaag gaagctaaaa
     2521 tggagaaaaa aatcactgga tataccaccg ttgatatatc ccaatggcat cgtaaagaac
     2581 attttgaggc atttcagtca gttgctcaat gtacctataa ccagaccgtt cagctggata
     2641 ttacggcctt tttaaagacc gtaaagaaaa ataagcacaa gttttatccg gcctttattc
     2701 acattcttgc ccgcctgatg aatgctcatc cggaatttcg tatggcaatg aaagacggtg
     2761 agctggtgat atgggatagt gttcaccctt gttacaccgt tttccatgag caaactgaaa
     2821 cgttttcatc gctctggagt gaataccacg acgatttccg gcagtttcta cacatatatt
     2881 cgcaagatgt ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg tttattgaga
     2941 atatgttttt cgtctcagcc aatccctggg tgagtttcac cagttttgat ttaaacgtgg
     3001 ccaatatgga caacttcttc gcccccgttt tcaccatggg caaatattat acgcaaggcg
     3061 acaaggtgct gatgccgctg gcgattcagg ttcatcatgc cgtttgtgat ggcttccatg
     3121 tcggcagaat gcttaatgaa ttacaacagt actgcgatga gtggcagggc ggggcgtaat
     3181 ttttttaagg cagttattgg tgcccttaaa cgcctggggt aatgactctc tagcttgagg
     3241 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg
     3301 tcggtgaacg ctctcctgag taggacaaat ccgccctcta gagctgcctc gcgcgtttcg
     3361 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     3421 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     3481 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc ttaactatgc
     3541 ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg
     3601 cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg
     3661 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     3721 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     3781 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     3841 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     3901 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     3961 atacctgt
//
