LOCUS       sequence-500603-        2967 bp    DNA     circular SYN 20-FEB-2026
DEFINITION  EGFP expressed under PL TetO1 promoter; part of BNeu AS12; colE1 
            origin of replication.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2967)
  AUTHORS   Bonnerjee D, Chakraborty S, Mukherjee B, Basu R, Paul A, Bagh S
  TITLE     Multicellular artificial neural network-type architectures 
            demonstrate computational problem solving.
  JOURNAL   Nat Chem Biol. 2024 Nov;20(11):1524-1534. doi: 
            10.1038/s41589-024-01711-4. Epub 2024 Sep 16.
  PUBMED    39285005
REFERENCE   2  (bases 1 to 2967)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Feb 20, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41589-024-01711-4"; journalName: "Nat Chem Biol"; date: 
            "2024-11"; volume: "20"; issue: "11"; pages: "1524-1534"
COMMENT     Sequence Label: pM2_PLTetO1_R1_EGFP
FEATURES             Location/Qualifiers
     source          1..2967
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(7..25,26..31,32..48,49..54,55..80)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    7..25
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    32..50
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             90..101
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             join(108..110,111..113,114..827)
                     /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(108..110,111..113,114..824)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(153..174)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(414..433)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     761..782
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      851..937
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     rep_origin      complement(1102..1690)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1182..1201)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1778..1872)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(1902..2693,2694..2762))
                     /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     2525..2544
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2763..2867)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2935..2953
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagtccc tatcagtgat agagattgac atccctatca gtgatagaga tactgagcac
       61 atcagcagga cgcactgacc gaattcatta aagaggagaa aggtaccatg gtgagcaagg
      121 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
      181 gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
      241 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc
      301 tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct
      361 tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg
      421 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
      481 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
      541 actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga
      601 acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc
      661 agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc
      721 agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg
      781 tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaaagc ggccgcgact
      841 ctagaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg
      901 ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg ccgccctaga cctagggcgt
      961 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     1021 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     1081 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     1141 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     1201 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     1261 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     1321 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     1381 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     1441 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     1501 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     1561 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     1621 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     1681 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     1741 ctcacgttaa gggattttgg tcatgactag tgcttggatt ctcaccaata aaaaacgccc
     1801 ggcggcaacc gagcgttctg aacaaatcca gatggagttc tgaggtcatt actggatcta
     1861 tcaacaggag tccaagcgag ctcgtaaact tggtctgaca gttaccaatg cttaatcagt
     1921 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     1981 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     2041 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     2101 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     2161 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     2221 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     2281 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     2341 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     2401 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     2461 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     2521 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     2581 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     2641 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     2701 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     2761 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     2821 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     2881 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     2941 cgtatcacga ggccctttcg tcttcac
//
