LOCUS       sequence-475177-        6536 bp    DNA     circular SYN 07-JUL-2025
DEFINITION  Express NanoLuc-PPP2R1A 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 6536)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 6536)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 7, 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-PPP2R1A
FEATURES             Location/Qualifiers
     source          1..6536
                     /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             1065..1577
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     regulatory      1597..1606
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(3479..3497)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3534..3655
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3545..3564)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3599..3618
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3900..4257
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3900..4091
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3923..3943)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4108..4243
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        4272..4319
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4351..5145
                     /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(4405..4424)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5015..5034
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5209..5257
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(5496..6084)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5576..5595)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6200..6483
                     /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 ggctagcgct caccatggtc ttcacactcg
     1081 aagatttcgt tggggactgg cgacagacag ccggctacaa cctggaccaa gtccttgaac
     1141 agggaggtgt gtccagtttg tttcagaatc tcggggtgtc cgtaactccg atccaaagga
     1201 ttgtcctgag cggtgaaaat gggctgaaga tcgacatcca tgtcatcatc ccgtatgaag
     1261 gtctgagcgg cgaccaaatg ggccagatcg aaaaaatttt taaggtggtg taccctgtgg
     1321 atgatcatca ctttaaggtg atcctgcact atggcacact ggtaatcgac ggggttacgc
     1381 cgaacatgat cgactatttc ggacggccgt atgaaggcat cgccgtgttc gacggcaaaa
     1441 agatcactgt aacagggacc ctgtggaacg gcaacaaaat tatcgacgag cgcctgatca
     1501 accccgacgg ctccctgctg ttccgagtaa ccatcaacgg agtgaccggc tggcggctgt
     1561 gcgaacgcat tctggcgggc tcgagcggcg cgatcgccat ggcggcggcc gacggcgacg
     1621 actcgctgta ccccatcgcg gtgctcatag acgaactccg caatgaggac gttcagcttc
     1681 gcctcaacag catcaagaag ctgtccacca tcgccttggc ccttggggtt gaaaggaccc
     1741 gaagtgagct tctgcctttc cttacagata ccatctatga tgaagatgag gtcctcctgg
     1801 ccctggcaga acagctggga accttcacta ccctggtggg aggcccagag tacgtgcact
     1861 gcctgctgcc accgctggag tcgctggcca cagtggagga gacagtggtg cgggacaagg
     1921 cagtggagtc cttacgggcc atctcacacg agcactcgcc ctctgacctg gaggcgcact
     1981 ttgtgccgct agtgaagcgg ctggcgggcg gcgactggtt cacctcccgc acctcggcct
     2041 gcggcctctt ctccgtctgc tacccccgag tgtccagtgc tgtgaaggcg gaacttcgac
     2101 agtacttccg gaacctgtgc tcagatgaca cccccatggt gcggcgggcc gcagcctcca
     2161 agctggggga gtttgccaag gtgctggagc tggacaacgt caagagtgag atcatcccca
     2221 tgttctccaa cctggcctct gacgagcagg actcggtgcg gctgctggcg gtggaggcgt
     2281 gcgtgaacat cgcccagctt ctgccccagg aggatctgga ggccctggtg atgcccactc
     2341 tgcgccaggc cgctgaagac aagtcctggc gcgtccgcta catggtggct gacaagttca
     2401 cagagctcca gaaagcagtg gggcctgaga tcaccaagac agacctggtc cctgccttcc
     2461 agaacctgat gaaagactgt gaggccgagg tgagggccgc agcctcccac aaggtcaaag
     2521 agttctgtga aaacctctca gctgactgtc gggagaatgt gatcatgtcc cagatcttgc
     2581 cctgcatcaa ggagctggtg tccgatgcca accaacatgt caagtctgcc ctggcctcag
     2641 tcatcatggg tctctctccc atcttgggca aagacaacac catcgagcac ctcttgcccc
     2701 tcttcctggc tcagctgaag gatgagtgcc ctgaggtacg gctgaacatc atctctaacc
     2761 tggactgtgt gaacgaggtg attggcatcc ggcagctgtc ccagtccctg ctccctgcca
     2821 ttgtggagct ggctgaggac gccaagtggc gggtgcggct ggccatcatt gagtacatgc
     2881 ccctcctggc tggacagctg ggagtggagt tctttgatga gaaacttaac tccttgtgca
     2941 tggcctggct tgtggatcat gtatatgcca tccgcgaggc agccaccagc aacctgaaga
     3001 agctagtgga aaagtttggg aaggagtggg cccatgccac aatcatcccc aaggtcttgg
     3061 ccatgtccgg agaccccaac tacctgcacc gcatgactac gctcttctgc atcaatgtgc
     3121 tgtctgaggt ctgtgggcag gacatcacca ccaagcacat gctacccacg gttctgcgca
     3181 tggctgggga cccggttgcc aatgtccgct tcaatgtggc caagtctctg cagaagatag
     3241 ggcccatcct ggacaacagc accttgcaga gtgaagtcaa gcccatccta gagaagctga
     3301 cccaggacca ggatgtggac gtcaaatact ttgcccagga ggctctgact gttctgtctc
     3361 tcgccgttta aacgaattcg ggctcggtac ccggggatcc tctagagtcg acctgcaggc
     3421 atgcaagctg atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccacc
     3481 gctgagcaat aactagcata accccttggg gcggccgctt cgagcagaca tgataagata
     3541 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga
     3601 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa
     3661 caacaattgc attcatttta tgtttcaggt tcagggggag atgtgggagg tttttttaag
     3721 caagtaaaac ctctacaaat gtggtaaaat cgaattttaa caaaatatta acgcttacaa
     3781 tttcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgcgg
     3841 atctgcgcag caccatggcc tgaaataacc tctgaaagag gaacttggtt aggtaccttc
     3901 tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     3961 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     4021 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     4081 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     4141 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     4201 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     4261 cttaattaac tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca
     4321 aggtgaggaa ctaaacccag gaggcagatc atgattgaac aagatggatt gcacgcaggt
     4381 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
     4441 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
     4501 accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg
     4561 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
     4621 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
     4681 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
     4741 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
     4801 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
     4861 ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
     4921 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
     4981 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
     5041 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
     5101 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
     5161 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg atggccgcaa taaaatatct
     5221 ttattttcat tacatctgtg tgttggtttt ttgtgtgaat cgatagcgat aaggatcctc
     5281 tttgcgcttg cgttttccct tgtccagata gcccagtagc tgacattcat ccggggtcag
     5341 caccgtttct gcggactggc tttctacccg gtatcagctc actcaaaggc ggtaatacgg
     5401 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     5461 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     5521 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     5581 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5641 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     5701 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     5761 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     5821 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     5881 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     5941 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6001 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6061 acgcgcagaa aaaaaggatt tcaagaagat cctttgatct tttctacggg gtctgacgct
     6121 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     6181 acctagatcc ttttatagtc cggaaataca ggaacgcacg ctggatggcc cttcgctggg
     6241 atggtgaaac catgaaaaat ggcagcttca gtggattaag tgggggtaat gtggcctgta
     6301 ccctctggtt gcataggtat tcatacggtt aaaatttatc aggcgcgatt gcggcagttt
     6361 ttcgggtggt ttgttgccat ttttacctgt ctgctgccgt gatcgcgctg aacgcgtttt
     6421 agcggtgcgt acaattaagg gattatggta aatccactta ctgtctgccc tcgtagccat
     6481 cgagataaac cgcagtactc cggccacgat gcgtccggcg tagaggatcg agatct
//
