LOCUS       sequence-296184-        4834 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the CeNLuc LumiFluor in bacterial cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4834)
  AUTHORS   Parag-Sharma K, O'Banion CP, Henry EC, Musicant AM, Cleveland JL, 
            Lawrence DS, Amelio AL
  TITLE     Engineered BRET-Based Biologic Light Sources Enable Spatio-Temporal 
            Control Over Diverse Optogenetic Systems.
  JOURNAL   ACS Synth Biol. 2019 Dec 13. doi: 10.1021/acssynbio.9b00277.
  PUBMED    31834783
REFERENCE   2  (bases 1 to 4834)
  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: "ACS Synth 
            Biol. 2019 Dec 13. doi: 10.1021/acssynbio.9b00277."
COMMENT     Sequence Label: pET20b-CeNLuc-6xHis
FEATURES             Location/Qualifiers
     source          1..4834
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             51..73
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             join(84..86,87..89,90..800)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAIHGNVYITADKQKNGIK
                     ANFGLNCNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(129..150)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(390..409)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     737..758
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             816..1328
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             1335..1352
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1405..1423)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1419..1466
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1503..1958
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1590..1609)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1800..1821
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1984..2088
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2089..2157,2158..2949)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2307..2326)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3120..3708
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3609..3628
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3862..3879
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3894..4033
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3959..3978)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4119..4141
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4135..4326)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 taatacgact cactataggg agaccacaac ggtttccctc tagaaataat tttgtttaac
       61 tttaagaagg agatatacat atgatggtga gcaagggcga ggagctgttc accggggtgg
      121 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
      181 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca
      241 agctgcccgt gccctggccc accctcgtga ccaccctgag ctggggcgtg cagtgcttcg
      301 cccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
      361 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
      421 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
      481 aggacggcaa catcctgggg cacaagctgg agtacaacgc catccacggc aacgtctata
      541 tcaccgccga caagcagaag aacggcatca aggccaactt cggcctcaac tgcaacatcg
      601 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
      661 ccgtgctgct gcccgacaac cactacctga gcacccagtc caagctgagc aaagacccca
      721 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
      781 gcatggacga gctgtacaag gatatctccg gaggtatggt cttcacactc gaagatttcg
      841 ttggggactg gcgacagaca gccggctaca acctggacca agtccttgaa cagggaggtg
      901 tgtccagttt gtttcagaat ctcggggtgt ccgtaactcc gatccaaagg attgtcctga
      961 gcggtgaaaa tgggctgaag atcgacatcc atgtcatcat cccgtatgaa ggtctgagcg
     1021 gcgaccaaat gggccagatc gaaaaaattt ttaaggtggt gtaccctgtg gatgatcatc
     1081 actttaaggt gatcctgcac tatggcacac tggtaatcga cggggttacg ccgaacatga
     1141 tcgactattt cggacggccg tatgaaggca tcgccgtgtt cgacggcaaa aagatcactg
     1201 taacagggac cctgtggaac ggcaacaaaa ttatcgacga gcgcctgatc aaccccgacg
     1261 gctccctgct gttccgagta accatcaacg gagtgaccgg ctggcggctg tgcgaacgca
     1321 ttctggcgct cgagcaccac caccaccacc actgagatcc ggctgctaac aaagcccgaa
     1381 aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct
     1441 ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga ttggcgaatg
     1501 ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     1561 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     1621 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     1681 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     1741 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     1801 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     1861 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     1921 taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat
     1981 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     2041 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     2101 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     2161 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     2221 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     2281 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     2341 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     2401 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     2461 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     2521 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     2581 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg
     2641 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     2701 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     2761 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     2821 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     2881 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     2941 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     3001 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     3061 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     3121 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     3181 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     3241 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     3301 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     3361 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     3421 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     3481 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     3541 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     3601 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     3661 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     3721 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     3781 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     3841 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
     3901 cggtattttc tccttacgca tctgtgcggt atttcacacc gcaatggtgc actctcagta
     3961 caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg
     4021 ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     4081 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     4141 gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc
     4201 gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag
     4261 aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt
     4321 ggtcactgat gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa
     4381 acgagagagg atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg
     4441 ttgtgagggt aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg
     4501 tcaatgccag cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc
     4561 tgcgatgcag atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta
     4621 cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca
     4681 gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc
     4741 ccgccagcct agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggccagga
     4801 cccaacgctg cccgagatct cgatcccgcg aaat
//
