LOCUS       sequence-270902-        4508 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 4508)
  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 4508)
  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-RF1
FEATURES             Location/Qualifiers
     source          1..4508
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(16..33)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(187..775)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(267..286)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(946..1737,1738..1806))
                     /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     1569..1588
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1807..1911)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1979..1997
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(2021..2035,2036..2041,2042..2058,2059..2065)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    2041..2057
                     /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    2073..2089
                     /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             2108..2119
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2138..2155
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      3266..3360
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             3404..4063
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3470..3489)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      4128..4214
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(4261..4283)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    join(4369..4508,1)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4424..4443
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
       61 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
      121 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
      181 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
      241 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
      301 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
      361 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
      421 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
      481 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
      541 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
      601 gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc
      661 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
      721 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
      781 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
      841 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
      901 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
      961 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     1021 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     1081 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     1141 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     1201 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     1261 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     1321 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     1381 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     1441 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     1501 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     1561 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     1621 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     1681 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     1741 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     1801 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     1861 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     1921 ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa
     1981 taggcgtatc acgaggccct ttcgtcttca cctcgagaaa tcataaaaaa tttatttgct
     2041 ttgtgagcgg ataacaatta taatagattc aattgtgagc ggataacaat ttcacacaga
     2101 attcattaaa gaggagaaat taactatgag aggatcgcat caccatcacc atcacggatc
     2161 catgaagcct tctatcgttg ccaaactgga agccctgcat gaacgccatg aagaagttca
     2221 ggcgttgctg ggtgacgcgc aaactatcgc cgaccaggaa cgttttcgcg cattatcacg
     2281 cgaatatgcg cagttaagtg atgtttcgcg ctgttttacc gactggcaac aggttcagga
     2341 agatatcgaa accgcacaga tgatgctcga tgatcctgaa atgcgtgaga tggcgcagga
     2401 tgaactgcgc gaagctaaag aaaaaagcga gcaactggaa cagcaattac aggttctgtt
     2461 actgccaaaa gatcctgatg acgaacgtaa cgccttcctc gaagtccgag ccggaaccgg
     2521 cggcgacgaa gcggcgctgt tcgcgggcga tctgttccgt atgtacagcc gttatgccga
     2581 agcccgccgc tggcgggtag aaatcatgag cgccagcgag ggtgaacatg gtggttataa
     2641 agagatcatc gccaaaatta gcggtgatgg tgtgtatggt cgtctgaaat ttgaatccgg
     2701 cggtcatcgc gtgcaacgtg ttcctgctac ggaatcgcag ggtcgtattc atacttctgc
     2761 ttgtaccgtt gcggtaatgc cagaactgcc tgacgcagaa ctgccggaca tcaacccagc
     2821 agatttacgc attgatactt tccgctcgtc aggggcgggt ggtcagcacg ttaacaccac
     2881 cgattcggca attcgtatta ctcacttgcc gaccgggatt gttgttgaat gtcaggacga
     2941 acgttcacaa cataaaaaca aagctaaagc actttctgtt ctcggtgctc gcatccacgc
     3001 tgctgaaatg gcaaaacgcc aacaggccga agcgtctacc cgtcgtaacc tgctggggag
     3061 tggcgatcgc agcgaccgta accgtactta caacttcccg caggggcgcg ttaccgatca
     3121 ccgcatcaac ctgacgctct accgcctgga tgaagtgatg gaaggtaagc tggatatgct
     3181 gattgaaccg attatccagg aacatcaggc cgaccaactg gcggcgttgt ccgagcagga
     3241 ataaaagctt aattagctga gcttggactc ctgttgatag atccagtaat gacctcagaa
     3301 ctccatctgg atttgttcag aacgctcggt tgccgccggg cgttttttat tggtgagaat
     3361 ccaagctagc ttggcgagat tttcaggagc taaggaagct aaaatggaga aaaaaatcac
     3421 tggatatacc accgttgata tatcccaatg gcatcgtaaa gaacattttg aggcatttca
     3481 gtcagttgct caatgtacct ataaccagac cgttcagctg gatattacgg cctttttaaa
     3541 gaccgtaaag aaaaataagc acaagtttta tccggccttt attcacattc ttgcccgcct
     3601 gatgaatgct catccggaat ttcgtatggc aatgaaagac ggtgagctgg tgatatggga
     3661 tagtgttcac ccttgttaca ccgttttcca tgagcaaact gaaacgtttt catcgctctg
     3721 gagtgaatac cacgacgatt tccggcagtt tctacacata tattcgcaag atgtggcgtg
     3781 ttacggtgaa aacctggcct atttccctaa agggtttatt gagaatatgt ttttcgtctc
     3841 agccaatccc tgggtgagtt tcaccagttt tgatttaaac gtggccaata tggacaactt
     3901 cttcgccccc gttttcacca tgggcaaata ttatacgcaa ggcgacaagg tgctgatgcc
     3961 gctggcgatt caggttcatc atgccgtttg tgatggcttc catgtcggca gaatgcttaa
     4021 tgaattacaa cagtactgcg atgagtggca gggcggggcg taattttttt aaggcagtta
     4081 ttggtgccct taaacgcctg gggtaatgac tctctagctt gaggcatcaa ataaaacgaa
     4141 aggctcagtc gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc
     4201 tgagtaggac aaatccgccc tctagagctg cctcgcgcgt ttcggtgatg acggtgaaaa
     4261 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     4321 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac
     4381 ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt
     4441 gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     4501 cgcatcag
//
