LOCUS       sequence-475173-        7027 bp    DNA     circular SYN 24-JUN-2025
DEFINITION  Express NanoLuc-PPP2R5A 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 7027)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 7027)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 24, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: NanoBRET Assay Vector, NanoLuc-PPP2R5A
FEATURES             Location/Qualifiers
     source          1..7027
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    17..98
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     42..61
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1780..2292)
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     promoter        complement(2331..2349)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2332..2349)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        2469..2772
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2927..2947
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        2980..3027
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3059..3718
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3125..3144)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     rep_origin      complement(3890..4478)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3970..3989)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4604..4652
                     /note="synthetic polyadenylation signal"
     CDS             complement(4676..5470)
                     /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(4787..4806)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5397..5416
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(5849..7027)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          5858..6796
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(5881..5901)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
ORIGIN
        1 aagacgcgtg acgtcggcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
       61 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
      121 ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
      181 aatgtggtat ggctgattat gatccgcggc cgcaatacgt cgacgttatc agctgacttc
      241 gtacgagagc ctaggtgttt ctagaggatc cccgggtacc gagcccgaat tcgtttaaac
      301 ttcggcactt gtattgctga gaatactgtg catgttgtaa gcactattct gtttttctag
      361 agctttcttt agctttagct cctctaattt tttccataat tcttcacgtt ccaattcctt
      421 ctttttctct ctctgtcttt cagctttgta tgagctagta aggtcatcga aaagcttgcc
      481 attcatttcc attagggttt tcagcacatt gtataccagt gctacaatgg tcggattcca
      541 gtgttctttg gaaattttgt acaaactggc aaacataatt ggcagaattt tatcaatgtt
      601 ctcctcaatc aaactaagaa tatattcgtt attccagaag tacaatgccc tttctgcaac
      661 ctgaaaatga gaactggata cacacttgga tatctgcttg aaaagtggct cttcaatttt
      721 tttgaactgt gttggttcaa tgacatctaa gatttcttca atttctccta aaaacatcac
      781 ctctttctga ctgcaggttt ttggccaaaa tttcagcagt cctctgatca ctggctctgt
      841 tagtgttgta tctttctcca ggaactgtac aacacaatat gctagctgag catgaaacaa
      901 agctaatcct tttgcagtat gcataggaat aagaaccttc attagaaatt gtttatgttc
      961 tgctttcagt ggcaatgcaa agccattgat aatacttcct aatatttcaa gaagttcagc
     1021 aacaccattg aaatgttctg tttcatatat aaacctgagg aaaatgttgt taatttgttt
     1081 tctgatgaat gctcttaatc caagaaattt cccataaatt cggtgcagaa cagtcttcag
     1141 gaagtcacgt tctctgggat cttcactatc aaaaagctcc aggagctgtt gtacgaattt
     1201 ctgatcaatg tatcgttttg caatgctagg ctggaaatca gggctctcca aaaatctcaa
     1261 gaagaattca tataccaact gtatgtgagg ccaagaggcc tcaagcgtgg gttcatcctc
     1321 ttctggatca aaatctggat tatcacttgg aggaagtgta cggaagatgt tagcactgat
     1381 catttttact atatcagaat acgctgattc aacaattaca ccacgattag ttgaaacata
     1441 ctcaaccagt tcattcagtg ttgctctttt aatttctttg ctcttcaagt ctgaaacaga
     1501 gtccatgaaa tcaaacagta tacaacactg ctgcaacttc tgacagaaaa gctcttgttg
     1561 ttcatttgaa gtggcatctt tgagctgggg cagcgggtgc agctctgcct ggctgccctg
     1621 gctgcgaaac tgcgacgagc cctgggagcg cttctgcctc tgcgccttgc ggaccgattt
     1681 ccgggtgaag ccgtccactt tctccgaggc cgagatggcg gcgctggcag cccccgccgg
     1741 cggcgacgac gacgacatgg cgatcgcgcc gctcgagccc gccagaatgc gttcgcacag
     1801 ccgccagccg gtcactccgt tgatggttac tcggaacagc agggagccgt cggggttgat
     1861 caggcgctcg tcgataattt tgttgccgtt ccacagggtc cctgttacag tgatcttttt
     1921 gccgtcgaac acggcgatgc cttcatacgg ccgtccgaaa tagtcgatca tgttcggcgt
     1981 aaccccgtcg attaccagtg tgccatagtg caggatcacc ttaaagtgat gatcatccac
     2041 agggtacacc accttaaaaa ttttttcgat ctggcccatt tggtcgccgc tcagaccttc
     2101 atacgggatg atgacatgga tgtcgatctt cagcccattt tcaccgctca ggacaatcct
     2161 ttggatcgga gttacggaca ccccgagatt ctgaaacaaa ctggacacac ctccctgttc
     2221 aaggacttgg tccaggttgt agccggctgt ctgtcgccag tccccaacga aatcttcgag
     2281 tgtgaagacc atggtgagcg ctagctgcaa aaagaacaag tagcttgtat tctatagtgt
     2341 cacctaaatg ggcgatctga cggttcacta aaccagctct gcttatatag gcctccctgg
     2401 cgcgccacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
     2461 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
     2521 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
     2581 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
     2641 catatgccaa gtccgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
     2701 gcccagtaca tgaccttacg ggactttcct acttggcagt acatctacgt attagtcatc
     2761 gctattacca tggtgatgcg gttttggcag tacaccaatg ggcgtggata gcggtttgac
     2821 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
     2881 aatcaacggg actttccaaa atgtcgtaat aaccccgccc cgttgacgca aatgggcggt
     2941 aggcgtgtac ggtgggaggt caaatcgaat tctaatactg ttgacaatta atcatcggca
     3001 tagtatatcg gcatagtata atacgacaag gtgaggaact aaacccagga ggcagatcat
     3061 ggagaaaaaa atcactggat ataccaccgt tgatatatcc caatggcatc gtaaagaaca
     3121 ttttgaggca tttcagtcag ttgctcaatg tacctataac cagaccgttc agctggatat
     3181 tacggccttt ttaaagaccg taaagaaaaa taagcacaag ttttatccgg cctttattca
     3241 cattcttgcc cgcctgatga atgctcatcc ggaattccgt atggcaatga aagacggtga
     3301 gctggtgata tgggatagtg ttcacccttg ttacaccgtt ttccatgagc aaactgaaac
     3361 gttttcatcg ctctggagtg aataccacga cgatttccgg cagtttctac acatatattc
     3421 gcaagatgtg gcgtgttacg gtgaaaacct ggcctatttc cctaaagggt ttattgagaa
     3481 tatgtttttc gtctcagcca atccctgggt gagtttcacc agttttgatt taaacgtggc
     3541 caatatggac aacttcttcg cccccgtttt cactatgggc aaatattata cgcaaggcga
     3601 caaggtgctg atgccgctgg cgattcaggt tcatcatgcc gtctgtgatg gcttccatgt
     3661 cggcagaatg cttaatgaat tacaacagta ctgcgatgag tggcagggcg gggcgtaagc
     3721 gggactctgg ggttcgaaat gaccgaccaa gcgacgccca accggtatca gctcactcaa
     3781 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     3841 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     3901 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     3961 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     4021 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     4081 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     4141 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     4201 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     4261 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     4321 acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     4381 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     4441 gcaagcagca gattacgcgc agaaaaaaag gatttcaaga agatcctttg atcttttcta
     4501 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     4561 caaaaaggat cttcacctag atccttttat agtccggaat cgtcacacaa aaaaccaaca
     4621 cacagatgta atgaaaataa agatatttta ttgcggccat cgtgatggct agagttcaga
     4681 agaactcgtc aagaaggcga tagaaggcga tgcgctgcga atcgggagcg gcgataccgt
     4741 aaagcacgag gaagcggtca gcccattcgc cgccaagctc ttcagcaata tcacgggtag
     4801 ccaacgctat gtcctgatag cggtccgcca cacccagccg gccacagtcg atgaatccag
     4861 aaaagcggcc attttccacc atgatattcg gcaagcaggc atcgccgtgg gtcacgacga
     4921 gatcctcgcc gtcgggcatg cgcgccttga gcctggcgaa cagttcggct ggcgcgagcc
     4981 cctgatgctc ttcgtccaga tcatcctgat cgacaagacc ggcttccatc cgagtacgtg
     5041 ctcgctcgat gcgatgtttc gcttggtggt cgaatgggca ggtagccgga tcaagcgtat
     5101 gcagccgccg cattgcatca gccatgatgg atactttctc ggcaggagca aggtgagatg
     5161 acaggagatc ctgccccggc acttcgccca atagcagcca gtcccttccc gcttcagtga
     5221 caacgtcgag cacagctgcg caaggaacgc ccgtcgtggc cagccacgat agccgcgctg
     5281 cctcgtcctg cagttcattc agggcaccgg acaggtcggt cttgacaaaa agaaccgggc
     5341 gcccctgcgc tgacagccgg aacacggcgg catcagagca gccgattgtc tgttgtgccc
     5401 agtcatagcc gaatagcctc tccacccaag cggccggaga acctgcgtgc aatccatctt
     5461 gttcaatcat ggtggcatcg attcccggtc aacatccagt acacctccgg caccaccggc
     5521 ttccccaagg gcgccacgct ctcccaccac aacatcctca acaacggcta ctgggtgggc
     5581 cgcacggtcg gctacacgga gcaggaccgg gtgtgtctgc cggtgccctt ctaccactgc
     5641 ttcggcatgg tcatgggcaa cttgggcgcc acctcccacg ccgcctgcat cgtcatcccg
     5701 gccccgtcct tcgagccggc ggcgacgctg gaggcggtgc agcgggagag gtgcacgtcc
     5761 ctgtacggcg tccccacgaa gccgaattcc accacactgg actagatgca tgctcgacag
     5821 tgtgatggat atctgcagaa ttcggctttc acgacacctg aaatggaaga aaaaaacttt
     5881 gaaccactgt ctgaggcttg agaatgaacc aagatccaaa ctcaaaaagg gcaaattcca
     5941 aggagaatta catcaagtgc caagctggcc taacttcagt ctccacccac tcagtgtggg
     6001 gaaactccat cgcataaaac ccctcccccc aacctaaaga cgacgtactc caaaagctcg
     6061 agaactaatc gaggtgcctg gacggcgccc ggtactccgt ggagtcacat gaagcgacgg
     6121 ctgaggacgg aaaggccctt ttcctttgtg tgggtgactc acccgcccgc tctcccgagc
     6181 gccgcgtcct ccattttgag ctgcctgcag cagggccggg aagcggccat ctttccgctc
     6241 acgcaactgg tgccgaccgg gccagccttg ccgcccaggg cggggcgata cacggcggcg
     6301 cgaggccagg caccagagca ggccggccag cttgagacta cccccgtccg attctcggtg
     6361 gccgcgctcg caggccccgc ctcgccgaac atgtgcgctg ggacgcacgg gccccgtcgc
     6421 cgcccgcggc cccaaaaacc gaaataccag tgtgcagatc ttggcccgca tttacaagac
     6481 tatcttgcca gaaaaaaagc gtcgcagcag gtcatcaaaa attttaaatg gctagagact
     6541 tatcgaaagc agcgagacag gcgcgaaggt gccaccagat tcgcacgcgg cggccccagc
     6601 gcccaggcca ggcctcaact caagcacgag gcgaaggggc tccttaagcg caaggcctcg
     6661 aactctccca cccacttcca acccgaagct cgggatcaag aatcacgtac tgcagccagg
     6721 tggaagtaat tcaaggcacg caagggccat aacccgtaaa gaggccaggc ccgcgggaac
     6781 cacacacggc acttacctgt gttctggcgg caaacccgtt gcgaaaaaga acgttcacgg
     6841 cgactactgc acttatatac ggttctcccc caccctcggg aaaaaggcgg agccagtaca
     6901 cgacatcact ttcccagttt accccgcgcc accttctcta ggcaccggtt caattgccga
     6961 cccctccccc caacttctcg gggactgtgg gcgatgtgcg ctctgcccac tgacgggcac
     7021 cggagcc
//
