LOCUS       sequence-474727-        6141 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express EED-NanoLuc(R) Fusion Protein in Mammalian Cells under a CMV
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6141)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6141)
  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: 
            "Unpublished"
COMMENT     Sequence Label: NanoBRET Assay Vector, EED-NanoLuc
FEATURES             Location/Qualifiers
     source          1..6141
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1033..1052
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1033..1051
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2484..2996
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(3084..3102)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3139..3260
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3150..3169)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3204..3223
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3505..3862
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3505..3696
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3528..3548)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3713..3848
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        3877..3924
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3956..4750
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4010..4029)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4620..4639
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4814..4862
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(5101..5689)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5181..5200)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    5805..6088
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tggtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt attaatacga ctcactatag ggctagcgat cgccatgtcc gagagggaag
     1081 tgtcgactgc gccggcggga acagacatgc ctgcggccaa gaagcagaag ctgagcagtg
     1141 acgagaacag caatccagac ctctctggag acgagaatga tgacgctgtc agtatagaaa
     1201 gtggtacaaa cactgaacgc cctgatacac ctacaaacac gccaaatgca cctggaagga
     1261 aaagttgggg aaagggaaaa tggaagtcaa agaaatgcaa atattctttc aaatgtgtaa
     1321 atagtctcaa ggaagatcat aaccaaccat tgtttggagt tcagtttaac tggcacagta
     1381 aagaaggaga tccattagtg tttgcaactg taggaagcaa cagagttacc ttgtatgaat
     1441 gtcattcaca aggagaaatc cggttgttgc aatcttacgt ggatgctgat gctgatgaaa
     1501 acttttacac ttgtgcatgg acctatgata gcaatacgag ccatcctctg ctggctgtag
     1561 ctggatctag aggcataatt aggataataa atcctataac aatgcagtgt ataaagcact
     1621 atgttggcca tggaaatgct atcaatgagc tgaaattcca tccaagagat ccaaatcttc
     1681 tcctgtcagt aagtaaagat catgctttac gattatggaa tatccagacg gacactctgg
     1741 tggcaatatt tggaggcgta gaagggcaca gagatgaagt tctaagtgct gattatgatc
     1801 ttttgggtga aaaaataatg tcctgtggta tggatcattc tcttaaactt tggaggatca
     1861 attcaaagag aatgatgaat gcaattaagg aatcttatga ttataatcca aataaaacta
     1921 acaggccatt tatttctcag aaaatccatt ttcctgattt ttctaccaga gacatacata
     1981 ggaattatgt tgattgtgtg cgatggttag gcgatttgat actttctaag agtggccgtg
     2041 ccattttaca ttcccaccag caatgtatga gagatccagt gtctccgaat cttcgccagc
     2101 atttgtcttg tgaaaatgcc attgtgtgct ggaaacctgg caagatggaa gatgatatag
     2161 ataaaattaa acccagtgaa tctaatgtga ctattcttgg gcgatttgat tacagccagt
     2221 gtgacatttg gtacatgagg ttttctatgg atttctggca aaagatgctt gcattgggca
     2281 atcaagttgg caaactttat gtttgggatt tagaagtaga agatcctcat aaagccaaat
     2341 gtacaacact gactcatcat aaatgtggtg ctgctattcg acaaaccagt tttagcaggg
     2401 atagcagcat tcttatagct gtttgtgatg atgccagtat ttggcgctgg gatcgacttc
     2461 gagtttctct cggctcgagc ggcgtcttca cactcgaaga tttcgttggg gactggcgac
     2521 agacagccgg ctacaacctg gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc
     2581 agaatctcgg ggtgtccgta actccgatcc aaaggattgt cctgagcggt gaaaatgggc
     2641 tgaagatcga catccatgtc atcatcccgt atgaaggtct gagcggcgac caaatgggcc
     2701 agatcgaaaa aatttttaag gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc
     2761 tgcactatgg cacactggta atcgacgggg ttacgccgaa catgatcgac tatttcggac
     2821 ggccgtatga aggcatcgcc gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt
     2881 ggaacggcaa caaaattatc gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc
     2941 gagtaaccat caacggagtg accggctggc ggctgtgcga acgcattctg gcgtaaggcc
     3001 gcgactctag agtcgacctg caggcatgca agctgatccg gctgctaaca aagcccgaaa
     3061 ggaagctgag ttggctgctg ccaccgctga gcaataacta gcataacccc ttggggcggc
     3121 cgcttcgagc agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc
     3181 agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta
     3241 taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg
     3301 gggagatgtg ggaggttttt ttaagcaagt aaaacctcta caaatgtggt aaaatcgaat
     3361 tttaacaaaa tattaacgct tacaatttcc tgatgcggta ttttctcctt acgcatctgt
     3421 gcggtatttc acaccgcata cgcggatctg cgcagcacca tggcctgaaa taacctctga
     3481 aagaggaact tggttaggta ccttctgagg cggaaagaac cagctgtgga atgtgtgtca
     3541 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3601 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3661 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     3721 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     3781 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     3841 tggaggccta ggcttttgca aaaagcttaa ttaactgttg acaattaatc atcggcatag
     3901 tatatcggca tagtataata cgacaaggtg aggaactaaa cccaggaggc agatcatgat
     3961 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     4021 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     4081 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     4141 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     4201 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     4261 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     4321 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     4381 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     4441 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     4501 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     4561 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     4621 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     4681 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     4741 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     4801 tcacgatggc cgcaataaaa tatctttatt ttcattacat ctgtgtgttg gttttttgtg
     4861 tgaatcgata gcgataagga tcctctttgc gcttgcgttt tcccttgtcc agatagccca
     4921 gtagctgaca ttcatccggg gtcagcaccg tttctgcgga ctggctttct acccggtatc
     4981 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     5041 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     5101 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     5161 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     5221 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     5281 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     5341 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     5401 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     5461 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     5521 taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac
     5581 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     5641 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatttcaag aagatccttt
     5701 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     5761 catgagatta tcaaaaagga tcttcaccta gatcctttta tagtccggaa atacaggaac
     5821 gcacgctgga tggcccttcg ctgggatggt gaaaccatga aaaatggcag cttcagtgga
     5881 ttaagtgggg gtaatgtggc ctgtaccctc tggttgcata ggtattcata cggttaaaat
     5941 ttatcaggcg cgattgcggc agtttttcgg gtggtttgtt gccattttta cctgtctgct
     6001 gccgtgatcg cgctgaacgc gttttagcgg tgcgtacaat taagggatta tggtaaatcc
     6061 acttactgtc tgccctcgta gccatcgaga taaaccgcag tactccggcc acgatgcgtc
     6121 cggcgtagag gatcgagatc t
//
