LOCUS       sequence-452405-        3238 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PrOR1OR2-driven expression of MGapt and deGFP for quantifying 
            cell-free RNA and protein synthesis..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3238)
  AUTHORS   Kapasiawala M, Murray RM
  TITLE     Metabolic perturbations to an E. coli-based cell-free system reveal 
            a trade-off between transcription and translation
  JOURNAL   bioRxiv 2023.03.22.533877
REFERENCE   2  (bases 1 to 3238)
  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: "bioRxiv 
            2023.03.22.533877"
COMMENT     Sequence Label: pBEST-PrOR1OR2-MGapt-UTR1-deGFP
FEATURES             Location/Qualifiers
     source          1..3238
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     6..24
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     RBS             398..420
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(457..478)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(718..737)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1065..1086
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1281..1298)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1452..2040)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1532..1551)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2211..3002,3003..3071))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2834..2853
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3072..3176)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ataaaaatag gcgtatcacg aggccctttc gtcttcaaga attctggcga atcctctgac
       61 cagccagaaa acgacctttc tgtggtgaaa ccggatgctg caattcagag cgccagcaag
      121 tgggggacag cagaagccct gaccgccgca gagtggatgt ttgacatggt gaagactatc
      181 gcaccatcag ccagaaaacc gaattttgct gggtgggcta acgatatccg cggatcccga
      241 ctgatgcgtg aacgtgacgg acgtaaccac cgcgacatgt gtgtgctgtt ctgagctaac
      301 accgtgcgtg ttgacaattt tacctctggc ggtgataatg gttgcagcta gcgggatccc
      361 gactggcgag agccaggtaa cgaatggatc caataatttt gtttaacttt aagaaggaga
      421 tataccatgg agcttttcac tggcgttgtt cccatcctgg tcgagctgga cggcgacgta
      481 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
      541 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
      601 accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac
      661 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
      721 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
      781 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
      841 tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag
      901 gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac
      961 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
     1021 acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
     1081 ttcgtgaccg ccgccgggat ctaactcgag caaagcccgc cgaaaggcgg gcttttctgt
     1141 gtcgaccgat gcccttgaga gccttcaacc cagtcagctc cttccggtgg gcgcggggca
     1201 tgactatcgt cgccgcactt atgactgtct tctttatcat gcaactcgta ggacaggtgc
     1261 cggcagcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     1321 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     1381 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     1441 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     1501 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     1561 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     1621 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     1681 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     1741 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     1801 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     1861 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     1921 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     1981 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     2041 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     2101 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     2161 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     2221 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     2281 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     2341 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     2401 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     2461 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     2521 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     2581 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     2641 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     2701 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     2761 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     2821 gcgtcaacac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     2881 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     2941 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     3001 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     3061 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     3121 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     3181 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacct
//
