LOCUS       sequence-63377-e        3293 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3293)
  AUTHORS   Shin J, Noireaux V
  TITLE     Efficient cell-free expression with the endogenous E. Coli RNA 
            polymerase and sigma factor 70.
  JOURNAL   J Biol Eng. 2010 Jun 24;4:8.
  PUBMED    20576148
REFERENCE   2  (bases 1 to 3293)
  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"; journalName: "J Biol 
            Eng."; date: "2010-06-24"; volume: "4"; pages: "8"
COMMENT     Sequence Label: pBEST-p15A-PLtetO1-deGFP-T500
FEATURES             Location/Qualifiers
     source          1..3293
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             7..29
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(66..87)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(327..346)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     674..695
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     rep_origin      1000..1544
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     1445..1464
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1661..1678
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(join(1911..2702,2703..2771))
                     /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     2534..2553
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2772..2876)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2944..2962
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     protein_bind    3233..3251
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3258..3276
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
ORIGIN
        1 aataattttg tttaacttta agaaggagat ataccatgga gcttttcact ggcgttgttc
       61 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
      121 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
      181 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
      241 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
      301 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
      361 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
      421 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
      481 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
      541 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
      601 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
      661 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc taactcgagc
      721 aaagcccgcc gaaaggcggg cttttctgtg tcgaccgatg cccttgagag ccttcaaccc
      781 agtcagctcc ttccggtggg cgcggggcat gactatcgtc gccgcactta tgactgtctt
      841 ctttatcatg caactcgtag gacaggtgcc ggcatgataa gctgtcaaac atgagaatta
      901 caacttatat cgtatggggc tgacttcagg tgctacattt gaagagataa attgcactga
      961 aatctagaaa tattttatct gattaataag atgatcttct tgagatcgtt ttggtctgcg
     1021 cgtaatctct tgctctgaaa acgaaaaaac cgccttgcag ggcggttttt cgaaggttct
     1081 ctgagctacc aactctttga accgaggtaa ctggcttgga ggagcgcagt caccaaaact
     1141 tgtcctttca gtttagcctt aaccggcgca tgacttcaag actaactcct ctaaatcaat
     1201 taccagtggc tgctgccagt ggtgcttttg catgtctttc cgggttggac tcaagacgat
     1261 agttaccgga taaggcgcag cggtcggact gaacgggggg ttcgtgcata cagtccagct
     1321 tggagcgaac tgcctacccg gaactgagtg tcaggcgtgg aatgagacaa acgcggccat
     1381 aacagcggaa tgacaccggt aaaccgaaag gcaggaacag gagagcgcac gagggagccg
     1441 ccaggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccaccac tgatttgagc
     1501 gtcagatttc gtgatgcttg tcaggggggc ggagcctatg gaaaaacggc tttgccgcgg
     1561 ccctctcact tccctgttaa gtatcttcct ggcatcttcc aggaaatctc cgccccgttc
     1621 gtaagccatt tccgctcgcc gcagtcgaac gaccgagcgt agcgagtcag tgagcgagga
     1681 agcggaatat atcctgtatc acatattctg ctgacgcacc ggtgcagcct tttttctcct
     1741 gccacatgaa gcacttcact gacaccctca tcagtgccaa catagtaagc cagtatacac
     1801 tccgctaggg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     1861 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     1921 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     1981 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     2041 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     2101 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     2161 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     2221 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     2281 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     2341 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     2401 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     2461 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     2521 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     2581 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     2641 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     2701 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     2761 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     2821 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     2881 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
     2941 aaaaataggc gtatcacgag gccctttcgt cttcaagaat tctggcgaat cctctgacca
     3001 gccagaaaac gacctttctg tggtgaaacc ggatgctgca attcagagcg gcagcaagtg
     3061 ggggacagca gaagacctga ccgccgcaga gtggatgttt gacatggtga agactatcgc
     3121 accatcagcc agaaaaccga attttgctgg gtgggctaac gatatccgcc tgatgcgtga
     3181 acgtgacgga cgtaaccacc gcgacatgtg tgtgctgttc cgctgggcat gctccctatc
     3241 agtgatagag attgacatcc ctatcagtga tagagatact gagcacagct agc
//
