LOCUS       sequence-258681-        6097 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 6097)
  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 6097)
  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-IF2
FEATURES             Location/Qualifiers
     source          1..6097
                     /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"
     terminator      4511..4605
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             4649..5308
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(4715..4734)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      5373..5459
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(5506..5528)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5614..5754
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5669..5688
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5769..5786)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(5940..6097,1..431))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6020..6039)
                     /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 cggatccatg acagatgtaa cgattaaaac gctggccgca gagcgacaga
     1861 cctccgtgga acgcctggta cagcaatttg ctgatgcagg tatccggaag tctgctgacg
     1921 actctgtgtc tgcacaagag aaacagactt tgattgacca cctgaatcag aaaaattcag
     1981 gcccggacaa attgacgctg caacgtaaaa cacgcagcac ccttaacatt cctggtaccg
     2041 gtggaaaaag caaatcggta caaatcgaag tccgcaagaa acgcaccttt gtgaaacgcg
     2101 atccgcaaga ggctgaacgc cttgcagcgg aagagcaagc gcagcgtgaa gcggaagagc
     2161 aagcccgtcg tgaggcagaa gaatcggcta aacgcgaggc gcaacaaaaa gctgaacgtg
     2221 aggccgcaga acaagctaag cgtgaagctg ctgaacaagc gaaacgtgaa gctgcggaaa
     2281 aagacaaagt gagcaatcaa caagacgata tgactaaaaa cgcccaggct gaaaaagccc
     2341 gccgtgagca ggaagctgca gagctcaagc gtaaagctga agaagaagcg cgtcgtaaac
     2401 tcgaagaaga agcacgtcgc gttgctgaag aagcacgtcg tatggcggaa gaaaacaaat
     2461 ggactgataa cgcggaaccg actgaagatt ccagcgatta tcacgtcact acttctcaac
     2521 atgctcgcca ggcagaagac gaaagcgatc gtgaagtcga aggcggccgt ggccgtggtc
     2581 gtaacgcgaa agcagcgcgt ccgaagaaag gcaacaaaca cgctgaatca aaagctgatc
     2641 gtgaagaagc acgcgcagca gtacgtggcg gtaaaggcgg aaaacgtaaa ggttcttcgc
     2701 tgcagcaagg cttccagaag cctgctcagg ccgttaaccg tgacgttgtg atcggcgaaa
     2761 ctatcaccgt tggcgaactg gcgaacaaga tggcggttaa aggctctcag gtcatcaaag
     2821 cgatgatgaa actgggcgca atggcaacca tcaaccaggt tatcgatcag gaaaccgcac
     2881 agctggttgc tgaagagatg ggccataaag ttatcctgcg tcgtgaaaac gagctggaag
     2941 aggcggtaat gagcgaccgt gacacgggtg ctgcggctga accgcgcgcg ccggttgtga
     3001 ccatcatggg tcacgttgac cacggtaaaa cctctctgct ggactacatt cgttcaacga
     3061 aagtggcctc tggcgaagcg ggcggcatta cccagcacat tggtgcatac cacgttgaaa
     3121 ctgaaaacgg catgatcacc ttcctggaca ccccggggca cgccgcgttt acttcaatgc
     3181 gtgctcgtgg tgcgcaggca acggacatcg tagtcctggt tgttgctgcc gacgacggtg
     3241 tgatgccgca gaccatcgaa gcaatccagc acgcgaaagc ggcgcaggta ccggtggtgg
     3301 ttgcagtgaa caagatcgat aaaccagaag ctgatccgga tcgcgttaag aacgaactct
     3361 cccagtacgg catcctgccg gaagagtggg gcggtgaaag ccagttcgta cacgtatctg
     3421 cgaaagcggg taccggtatc gatgaactgc tggacgctat cctgctgcag gcggaagttc
     3481 tggagctgaa agcggtacgt aaaggtatgg cgagcggtgc ggttatcgaa tccttcctcg
     3541 ataaaggtcg tggtccggtt gctaccgttc tggtacgtga aggtactctg cacaagggcg
     3601 atatcgttct gtgtggcttc gaatacggtc gtgttcgtgc gatgcgtaac gaactgggtc
     3661 aggaagtgct ggaagcgggt ccgtccattc cggtggaaat cctcggcctg tccggcgtac
     3721 cggctgcggg tgatgaagtt accgttgtac gtgacgagaa gaaagcgcgt gaagttgcac
     3781 tctatcgtca gggtaaattc cgcgaagtta aactggcgcg tcagcagaaa tctaaactcg
     3841 agaacatgtt cgccaacatg accgaaggcg aagttcacga agtgaatatc gtcctgaagg
     3901 cagacgtaca gggttctgtc gaagcgatct ccgactcctt gctgaaactg tctactgacg
     3961 aagttaaagt gaagatcatc ggttctggcg taggtggtat caccgaaacc gacgccaccc
     4021 tggctgcggc gtccaacgcc atcctggttg gctttaacgt acgtgctgat gcctctgcac
     4081 gtaaagtgat tgaagcggaa agcctggatc tgcgttacta ctccgtcatc tataacctga
     4141 ttgacgaagt gaaagcggcg atgagcggta tgctgtctcc ggaactgaaa cagcagatta
     4201 tcggtctggc ggaagttcgt gacgtgttca aatcgccgaa atttggtgcc atcgcaggct
     4261 gtatggttac cgaaggtgtg gttaaacgtc acaacccgat ccgcgttctg cgtgacaacg
     4321 tggttatcta cgaaggcgag ctggagtccc tgcgccgctt caaagatgac gttaacgaag
     4381 tccgtaacgg tatggaatgt ggtatcggcg ttaagaacta caacgacgtc cgcactggcg
     4441 atgtgatcga agtattcgaa atcatcgaga tccaacgtac cattgcttaa agcttaatta
     4501 gctgagcttg gactcctgtt gatagatcca gtaatgacct cagaactcca tctggatttg
     4561 ttcagaacgc tcggttgccg ccgggcgttt tttattggtg agaatccaag ctagcttggc
     4621 gagattttca ggagctaagg aagctaaaat ggagaaaaaa atcactggat ataccaccgt
     4681 tgatatatcc caatggcatc gtaaagaaca ttttgaggca tttcagtcag ttgctcaatg
     4741 tacctataac cagaccgttc agctggatat tacggccttt ttaaagaccg taaagaaaaa
     4801 taagcacaag ttttatccgg cctttattca cattcttgcc cgcctgatga atgctcatcc
     4861 ggaatttcgt atggcaatga aagacggtga gctggtgata tgggatagtg ttcacccttg
     4921 ttacaccgtt ttccatgagc aaactgaaac gttttcatcg ctctggagtg aataccacga
     4981 cgatttccgg cagtttctac acatatattc gcaagatgtg gcgtgttacg gtgaaaacct
     5041 ggcctatttc cctaaagggt ttattgagaa tatgtttttc gtctcagcca atccctgggt
     5101 gagtttcacc agttttgatt taaacgtggc caatatggac aacttcttcg cccccgtttt
     5161 caccatgggc aaatattata cgcaaggcga caaggtgctg atgccgctgg cgattcaggt
     5221 tcatcatgcc gtttgtgatg gcttccatgt cggcagaatg cttaatgaat tacaacagta
     5281 ctgcgatgag tggcagggcg gggcgtaatt tttttaaggc agttattggt gcccttaaac
     5341 gcctggggta atgactctct agcttgaggc atcaaataaa acgaaaggct cagtcgaaag
     5401 actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc
     5461 cgccctctag agctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     5521 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     5581 gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga
     5641 tagcggagtg tatactggct taactatgcg gcatcagagc agattgtact gagagtgcac
     5701 catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgctct
     5761 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     5821 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     5881 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     5941 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     6001 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     6061 tctcctgttc cgaccctgcc gcttaccgga tacctgt
//
