LOCUS       sequence-389815-        3102 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses EGFP under the control of PR promoter, pUC origin of 
            replication, Ampicillin selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3102)
  AUTHORS   Sarkar K, Mukhopadhyay S, Bonnerjee D, Srivastava R, Bagh S
  TITLE     A frame-shifted gene, which rescued its function by non-natural 
            start codons and its application in constructing synthetic gene 
            circuits.
  JOURNAL   J Biol Eng. 2019 Mar 1;13:20. doi: 10.1186/s13036-019-0151-x. 
            eCollection 2019.
  PUBMED    30867677
REFERENCE   2  (bases 1 to 3102)
  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: "2019-03-1"; volume: "13:20. doi"; pages: " 
            10.1186/s13036-019-0151-x. eCollection 2019"
COMMENT     Sequence Label: pRA1EGFP
FEATURES             Location/Qualifiers
     source          1..3102
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             71..82
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             join(89..91,92..94,95..808)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(89..91,92..94,95..805)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(134..155)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(395..414)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     742..763
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      820..906
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     rep_origin      complement(1007..1595)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1087..1106)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1913..2007)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(2037..2828,2829..2897))
                     /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     2660..2679
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2898..3002)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3070..3088
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagtaac accgtgcgtg ttgactattt tacctctggc ggtgataatg gttgcatgta
       61 cgaattcatt aaagaggaga aaggtaccat ggtgagcaag ggcgaggagc tgttcaccgg
      121 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
      181 cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
      241 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg
      301 cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
      361 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
      421 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
      481 caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt
      541 ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa
      601 catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga
      661 cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga
      721 ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac
      781 tctcggcatg gacgagctgt acaagtaatc tagaggcatc aaataaaacg aaaggctcag
      841 tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct cctgagtagg
      901 acaaatccgc cgccctagac ctagtagggg ataacgcagg aaagaacatg tgagcaaaag
      961 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     1021 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     1081 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     1141 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     1201 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     1261 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     1321 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     1381 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     1441 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     1501 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     1561 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     1621 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     1681 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     1741 tatatgagta acctgaggct atggcagggc ctgccgcccc gacgttggct gcgagccctg
     1801 ggccttcacc cgaacttggg gggtggggtg gggaaaagga agaaacgcgg gcgtattggc
     1861 cccaatgggg tctcggtggg gtatcgacag agtgccagcc cctagtgctt ggattctcac
     1921 caataaaaaa cgcccggcgg caaccgagcg ttctgaacaa atccagatgg agttctgagg
     1981 tcattactgg atctatcaac aggagtccaa gcgagctcgt aaacttggtc tgacagttac
     2041 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     2101 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     2161 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     2221 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     2281 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     2341 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     2401 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     2461 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     2521 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     2581 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     2641 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     2701 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     2761 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     2821 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     2881 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     2941 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     3001 cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta
     3061 acctataaaa ataggcgtat cacgaggccc tttcgtcttc ac
//
