LOCUS       sequence-478496-        6422 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Express PPM1D-NanoLuc 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 6422)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6422)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 7, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: PPM1D-NanoLuc Fusion Vector
FEATURES             Location/Qualifiers
     source          1..6422
                     /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             2901..3413
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(3501..3519)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3556..3677
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3567..3586)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3621..3640
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3922..4279
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3922..4113
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3945..3965)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4130..4265
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        4294..4341
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4373..5167
                     /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(4427..4446)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5037..5056
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5231..5279
                     /note="synthetic polyadenylation signal"
     rep_origin      5436..5981
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     5882..5901
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6098..6115
                     /label=L4440
                     /note="L4440 vector, forward primer"
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 cgccatggcg gggctgtact
     1081 cgctgggagt gagcgtcttc tccgaccagg gcgggaggaa gtacatggag gacgttactc
     1141 aaatcgttgt ggagcccgaa ccgacggctg aagaaaagcc ctcgccgcgg cggtcgctgt
     1201 ctcagccgtt gcctccgcgg ccgtcgccgg ccgcccttcc cggcggcgaa gtctcgggga
     1261 aaggcccagc ggtggcagcc cgagaggctc gcgaccctct cccggacgcc ggggcctcgc
     1321 cggcacctag ccgctgctgc cgccgccgtt cctccgtggc ctttttcgcc gtgtgcgacg
     1381 ggcacggcgg gcgggaggcg gcacagtttg cccgggagca cttgtggggt ttcatcaaga
     1441 agcagaaggg tttcacctcg tccgagccgg ctaaggtttg cgctgccatc cgcaaaggct
     1501 ttctcgcttg tcaccttgcc atgtggaaga aactggcgga atggccaaag actatgacgg
     1561 gtcttcctag cacatcaggg acaactgcca gtgtggtcat cattcggggc atgaagatgt
     1621 atgtagctca cgtaggtgac tcaggggtgg ttcttggaat tcaggatgac ccgaaggatg
     1681 actttgtcag agctgtggag gtgacacagg accataagcc agaacttccc aaggaaagag
     1741 aacgaatcga aggacttggt gggagtgtaa tgaacaagtc tggggtgaat cgtgtagttt
     1801 ggaaacgacc tcgactcact cacaatggac ctgttagaag gagcacagtt attgaccaga
     1861 ttccttttct ggcagtagca agagcacttg gtgatttgtg gagctatgat ttcttcagtg
     1921 gtgaatttgt ggtgtcacct gaaccagaca caagtgtcca cactcttgac cctcagaagc
     1981 acaagtatat tatattgggg agtgatggac tttggaatat gattccacca caagatgcca
     2041 tctcaatgtg ccaggaccaa gaggagaaaa aatacctgat gggtgagcat ggacaatctt
     2101 gtgccaaaat gcttgtgaat cgagcattgg gccgctggag gcagcgtatg ctccgagcag
     2161 ataacactag tgccatagta atctgcatct ctccagaagt ggacaatcag ggaaacttta
     2221 ccaatgaaga tgagttatac ctgaacctga ctgacagccc ttcctataat agtcaagaaa
     2281 cctgtgtgat gactccttcc ccatgttcta caccaccagt caagtcactg gaggaggatc
     2341 catggccaag ggtgaattct aaggaccata tacctgccct ggttcgtagc aatgccttct
     2401 cagagaattt tttagaggtt tcagctgaga tagctcgaga gaatgtccaa ggtgtagtca
     2461 taccctcaaa agatccagaa ccacttgaag aaaattgcgc taaagccctg actttaagga
     2521 tacatgattc tttgaataat agccttccaa ttggccttgt gcctactaat tcaacaaaca
     2581 ctgtcatgga ccaaaaaaat ttgaagatgt caactcctgg ccaaatgaaa gcccaagaaa
     2641 ttgaaagaac ccctccaaca aactttaaaa ggacattaga agagtccaat tctggccccc
     2701 tgatgaagaa gcatagacga aatggcttaa gtcgaagtag tggtgctcag cctgcaagtc
     2761 tccccacaac ctcacagcga aagaactctg ttaaactcac catgcgacgc agacttaggg
     2821 gccagaagaa aattggaaat cctttacttc atcaacacag gaaaactgtt tgtgtttgcg
     2881 tttctctcgg ctcgagcggc gtcttcacac tcgaagattt cgttggggac tggcgacaga
     2941 cagccggcta caacctggac caagtccttg aacagggagg tgtgtccagt ttgtttcaga
     3001 atctcggggt gtccgtaact ccgatccaaa ggattgtcct gagcggtgaa aatgggctga
     3061 agatcgacat ccatgtcatc atcccgtatg aaggtctgag cggcgaccaa atgggccaga
     3121 tcgaaaaaat ttttaaggtg gtgtaccctg tggatgatca tcactttaag gtgatcctgc
     3181 actatggcac actggtaatc gacggggtta cgccgaacat gatcgactat ttcggacggc
     3241 cgtatgaagg catcgccgtg ttcgacggca aaaagatcac tgtaacaggg accctgtgga
     3301 acggcaacaa aattatcgac gagcgcctga tcaaccccga cggctccctg ctgttccgag
     3361 taaccatcaa cggagtgacc ggctggcggc tgtgcgaacg cattctggcg taaggccgcg
     3421 actctagagt cgacctgcag gcatgcaagc tgatccggct gctaacaaag cccgaaagga
     3481 agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg gggcggccgc
     3541 ttcgagcaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
     3601 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
     3661 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
     3721 agatgtggga ggttttttta agcaagtaaa acctctacaa atgtggtaaa atcgaatttt
     3781 aacaaaatat taacgcttac aatttcctga tgcggtattt tctccttacg catctgtgcg
     3841 gtatttcaca ccgcatacgc ggatctgcgc agcaccatgg cctgaaataa cctctgaaag
     3901 aggaacttgg ttaggtacct tctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt
     3961 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     4021 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     4081 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     4141 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     4201 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     4261 aggcctaggc ttttgcaaaa agcttaatta actgttgaca attaatcatc ggcatagtat
     4321 atcggcatag tataatacga caaggtgagg aactaaaccc aggaggcaga tcatgattga
     4381 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     4441 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     4501 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga
     4561 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     4621 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     4681 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     4741 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     4801 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     4861 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga
     4921 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     4981 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     5041 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     5101 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     5161 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     5221 cgatggccgc aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga
     5281 atcgatagcg ataaggatcc gctcatgcga tgataagctg tcaaacatga gaattacaac
     5341 ttatatcgta tggggctgac ttcaggtgct acatttgaag agataaattg cactgaaatc
     5401 tagaaatatt ttatctgatt aataagatga tcttcttgag atcgttttgg tctgcgcgta
     5461 atctcttgct ctgaaaacga aaaaaccgcc ttgcagggcg gtttttcgaa ggttctctga
     5521 gctaccaact ctttgaaccg aggtaactgg cttggaggag cgcagtcacc aaaacttgtc
     5581 ctttcagttt agccttaacc ggcgcatgac ttcaagacta actcctctaa atcaattacc
     5641 agtggctgct gccagtggtg cttttgcatg tctttccggg ttggactcaa gacgatagtt
     5701 accggataag gcgcagcggt cggactgaac ggggggttcg tgcatacagt ccagcttgga
     5761 gcgaactgcc tacccggaac tgagtgtcag gcgtggaatg agacaaacgc ggccataaca
     5821 gcggaatgac accggtaaac cgaaaggcag gaacaggaga gcgcacgagg gagccgccag
     5881 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccaccactga tttgagcgtc
     5941 agatttcgtg atgcttgtca ggggggcgga gcctatggaa aaacggcttt gccgcggccc
     6001 tctcacttcc ctgttaagta tcttcctggc atcttccagg aaatctccgc cccgttcgta
     6061 agccatttcc gctcgccgca gtcgaacgac cgagcgtagc gagtcagtga gcgaggaagc
     6121 ggaatatatc ctgtatcaca tattctgctg acgcaccggt gcagcctttt ttctcctgcc
     6181 acatgaagca cttcactgac accctcatca gtgccaacat agtaagccag tatacactcc
     6241 gctagcgctg atgtccggcg gtgcttttgc cgttacgcac caccccgtca tgagattacg
     6301 cgttttagcg gtgcgtacaa ttaagggatt atggtaaatc cacttactgt ctgccctcgt
     6361 agccatcgag ataaaccgca gtactccggc cacgatgcgt ccggcgtaga ggatcgagat
     6421 ct
//
