LOCUS       sequence-275320-        4826 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 4826)
  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 4826)
  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-AsnRS
FEATURES             Location/Qualifiers
     source          1..4826
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(115..703)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(195..214)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(874..1665,1666..1734))
                     /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     1497..1516
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1735..1839)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1907..1925
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(1949..1963,1964..1969,1970..1986,1987..1993)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    1969..1985
                     /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    2001..2017
                     /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             2036..2047
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2066..2083
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      3512..3606
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             3650..4309
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3716..3735)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      4374..4460
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(4507..4529)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4615..4755
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4670..4689
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4770..4787)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
       61 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
      121 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
      181 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
      241 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
      301 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
      361 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
      421 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
      481 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
      541 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
      601 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
      661 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
      721 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
      781 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
      841 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
      901 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
      961 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     1021 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     1081 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     1141 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     1201 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     1261 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     1321 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     1381 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     1441 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     1501 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     1561 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     1621 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     1681 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     1741 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     1801 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     1861 acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac
     1921 gaggcccttt cgtcttcacc tcgagaaatc ataaaaaatt tatttgcttt gtgagcggat
     1981 aacaattata atagattcaa ttgtgagcgg ataacaattt cacacagaat tcattaaaga
     2041 ggagaaatta actatgagag gatcgcatca ccatcaccat cacggatcca tgagcgttgt
     2101 gcctgtagcc gacgtactcc agggccgtgt agccgttgac agcgaagtca ccgtgcgcgg
     2161 atgggtacgt acccgccgag attcaaaagc tggcatctcc ttcctcgccg tttatgacgg
     2221 ttcctgcttt gatcctgtac aggctgtcat caataattct ctgcccaatt acaatgaaga
     2281 cgtcctgcgt ctgaccaccg gctgctcggt cattgtgacg ggtaaagtcg tggcgtcgcc
     2341 gggccagggg caacaatttg aaattcaggc cagcaaggtt gaagttgctg gttgggttga
     2401 agatccagac acttacccga tggcggcaaa acgccacagc attgagtatc tgcgtgaagt
     2461 cgctcacctg cgtccgcgca caaacctgat tggtgccgtc gcgcgcgttc gccatacgct
     2521 ggcgcaggcg ctgcatcgct tctttaacga gcagggattc ttctgggttt caacgccact
     2581 gattaccgca tctgataccg aaggtgcagg cgaaatgttc cgcgtttcta cgctggatct
     2641 ggaaaacctg ccgcgtaacg atcagggcaa agtggatttc gacaaagact tctttggtaa
     2701 agagtctttc ctgaccgtat ctggccagtt gaacggcgaa acctacgctt gcgcattgtc
     2761 caaaatttat accttcggcc cgactttccg tgctgaaaac tccaacacca gccgtcacct
     2821 ggcggaattc tggatgctgg agccggaagt ggcgtttgct aacctgaacg atattgcggg
     2881 tctggctgaa gccatgctga aatatgtctt caaagcggtt ctcgaagaac gcgctgacga
     2941 catgaaattc ttcgctgaac gcgtagataa agatgccgtt tcacgtctgg aacgcttcat
     3001 tgaagccgat tttgcgcagg tggattatac cgacgcagtg accattctcg aaaactgcgg
     3061 caggaagttt gaaaacccgg tttactgggg agtcgatctc tcttctgagc atgagcgtta
     3121 tctggcggaa gaacacttta aagcaccggt agtggttaaa aactatccga aagatattaa
     3181 agcgttctat atgcgcctta acgaagacgg taaaaccgtt gcggctatgg acgttctggc
     3241 tccgggcatc ggtgagatca ttggtggctc ccagcgtgaa gaacgtctgg acgtgctgga
     3301 cgagcgtatg ctggaaatgg gcctgaataa agaagattac tggtggtatc gcgatctgcg
     3361 tcgctacggt actgttccgc attcaggttt cggtcttggt tttgaacgtc tgattgctta
     3421 cgtaactggc gtgcaaaacg tacgtgatgt gattccgttc ccacgtactc cgcgtaacgc
     3481 cagcttctaa aagcttaatt agctgagctt ggactcctgt tgatagatcc agtaatgacc
     3541 tcagaactcc atctggattt gttcagaacg ctcggttgcc gccgggcgtt ttttattggt
     3601 gagaatccaa gctagcttgg cgagattttc aggagctaag gaagctaaaa tggagaaaaa
     3661 aatcactgga tataccaccg ttgatatatc ccaatggcat cgtaaagaac attttgaggc
     3721 atttcagtca gttgctcaat gtacctataa ccagaccgtt cagctggata ttacggcctt
     3781 tttaaagacc gtaaagaaaa ataagcacaa gttttatccg gcctttattc acattcttgc
     3841 ccgcctgatg aatgctcatc cggaatttcg tatggcaatg aaagacggtg agctggtgat
     3901 atgggatagt gttcaccctt gttacaccgt tttccatgag caaactgaaa cgttttcatc
     3961 gctctggagt gaataccacg acgatttccg gcagtttcta cacatatatt cgcaagatgt
     4021 ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg tttattgaga atatgttttt
     4081 cgtctcagcc aatccctggg tgagtttcac cagttttgat ttaaacgtgg ccaatatgga
     4141 caacttcttc gcccccgttt tcaccatggg caaatattat acgcaaggcg acaaggtgct
     4201 gatgccgctg gcgattcagg ttcatcatgc cgtttgtgat ggcttccatg tcggcagaat
     4261 gcttaatgaa ttacaacagt actgcgatga gtggcagggc ggggcgtaat ttttttaagg
     4321 cagttattgg tgcccttaaa cgcctggggt aatgactctc tagcttgagg catcaaataa
     4381 aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg
     4441 ctctcctgag taggacaaat ccgccctcta gagctgcctc gcgcgtttcg gtgatgacgg
     4501 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     4561 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc
     4621 catgacccag tcacgtagcg atagcggagt gtatactggc ttaactatgc ggcatcagag
     4681 cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga
     4741 aaataccgca tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     4801 cggctgcggc gagcggtatc agctca
//
