LOCUS       sequence-479908-        8175 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Express EPOR-LgBiT/EPOR-SmBiT Fusion Proteins in Mammalian Cells 
            under a CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8175)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8175)
  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            "Unpublished"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: EPOR-LgBiT/EPOR-SmBiT Bidirectional Vector
FEATURES             Location/Qualifiers
     source          1..8175
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    24..105
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        complement(2375..2393)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2376..2393)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        2513..2816
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2817..3020
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2971..2991
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          3156..3288
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     3271..3290
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     3332..3351
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3332..3350
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    5064..5185
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5075..5094)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(5300..5333)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        5343..5390
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5422..6216
                     /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(5476..5495)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6086..6105
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      6433..6978
                     /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     6879..6898
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7095..7112
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    7262..7310
                     /note="synthetic polyadenylation signal"
     CDS             complement(7334..7732)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     regulatory      7729..7738
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(7763..8120)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7777..7912
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        7929..8120
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     8077..8097
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
ORIGIN
        1 aagacgcgtt aattaagacg tcggcagtga aaaaaatgct ttatttgtga aatttgtgat
       61 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
      121 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac
      181 ctctacaaat gtggtatggc tgattatgat ccgcggccgc aatacgtcga cgttatcagc
      241 tgacttcgta cgagagccta ggtgtttcta gagtcgcggc cttagctgtt gatggttact
      301 cggaacagca tggagccgtc gggggtgatc aggcgctcgt cgataatttt gttgccgttc
      361 cacagggtcc ctgttacagt gatctttttg ccgtcgaaca cggcgatgcc ttcatacggc
      421 cgtccgaaat agttcagcat gttcggcgta accccgtcga ttaccagtgt gccatagggc
      481 aggatcacct taaagtgatg atcatccaca gggtacacca ccttaaacac ctcttcgatc
      541 tgggccattt ggtcggcgct cagaccttca tacgggatga tgacatggat gtcgatcttc
      601 agggcatttt caccgctccg gacaatcctt tggatcggag ttacggacac ggcgagattc
      661 tgcagcaaac tggacacacc tccctgttca aggacttggt ccaggttgta ggcggctgtc
      721 tgttcccagt ccccaacgaa atcttcgagt gtgaagacac cgctcgagcc tccacctccg
      781 ctcccgccac cacctgaact gccttgagaa acagagcaag ccacatagct ggggggcaga
      841 ggctcagcgg ctgggataag gctgttctca taagggttgg agtaggggcc atcggataag
      901 cccccttggg ctccctggga gtcccctgag ctgtagtcag ttgagatgcc agagtcagat
      961 accacaaggt acaggtactt taggtggggt ggggtagggg gcagctcagg gcacagtgtc
     1021 catggacgca agagctggga gctggggtcc aggatagtgt actcaaagct ggcagcagag
     1081 gctccctctg ggctgggctt cgaggccaaa gcagatgagc aggaggatgc ttctgagcct
     1141 tcatccatgg ccactatgtc cacactgcca ccaggccctg ggaggtcctc actgggcggg
     1201 ttccggggca gcaaccattt gtccagcacc agataggtat cctgggcatg ctcactgccc
     1261 actggctcca gcagggggcc ctcatcatct gtccccggct ccactgcctg catcgtcccc
     1321 cagcagcgct ctgagaggac ttccagggaa gcaggtgggt cctccgtgaa gggggtgcag
     1381 gggctccacc acaggcagcc atcattctgg tacagccaca gctggaagtt acccttgtgg
     1441 gtggtgaaga ggccttcaaa ctcgctctct gggctcggga tgccaggcca gatcttctgc
     1501 ttcagagccc ggcggtggga gagcagcgcg agcacggtca gcagcaccag gatgaccacg
     1561 aggatgaggg agagcgtcag gatgaggggg tccaggtcgc taggcgtcag cagcgacaca
     1621 ggctccgacc aggcgctcca gaagccgccg aagctcggct cagccatacg cgcgcggacg
     1681 gcgaaggtgt agcgcgtccg gccccgcagg ttgctcagca cacactcggt gcggccctcc
     1741 aggatctcca ccctctgtac gctccctgcg ccgttgccgg ccgagacgtc cacctcgtag
     1801 cggatgtgag acgtcatggg tgtctcaggc ggcgggagcc agcgcaacac tacgtggccg
     1861 ctctcgtcag ccaaccgcgc caccagcccc acgggggcgt ctaggagcac tacttcattg
     1921 atgtggatga cacggtgata tcgcggagcg ccggaggctg ctgtgacgcg caactctagg
     1981 ggcacgaagc tcgacgtgtc ggctgtaggc agcgaacacc agaagcgcac cgcaccacga
     2041 gccgtgggag cctggtgcag gcgacacagc ttccatggct catcctcgag ctggtaggag
     2101 aagctgtagt tgcccgggcc caccccagcg ctcgccgctt cctcccagaa acacaccaag
     2161 tcctccaacc gctcggtgaa gcacagaagc tcttcgggcc cccgggccgc cagcaaggcc
     2221 gctttgctct cgaacttggg gtccgggagg ttaggcgggg gcgcccaggc ggccccagcg
     2281 agcaggagac aaagggagcc gacctggggc cagagggacg ccccgaggtg gtccatggcg
     2341 atcgctagct gcaaaaagaa caagtagctt gtattctata gtgtcaccta aatgggcgat
     2401 ctgacggttc actaaaccag ctctgcttat ataggcctcc ctggcgcgcc actagttatt
     2461 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
     2521 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
     2581 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
     2641 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtccgc
     2701 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
     2761 tacgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga
     2821 tgcggttttg gcagtacacc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
     2881 gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc
     2941 caaaatgtcg taataacccc gccccgttga cgcaaatggg cggtaggcgt gtacggtggg
     3001 aggtctatat aagcagagct ggtttagtga accgtcagat cactagaagc tttattgcgg
     3061 tagtttatca cagttaaatt gctaacgcag tcagtgcttc tgacacaaca gtctcgaact
     3121 taagctgcag aagttggtcg tgaggcactg ggcaggtaag tatcaaggtt acaagacagg
     3181 tttaaggaga ccaatagaaa ctgggcttgt cgagacagag aagactcttg cgtttctgat
     3241 aggcacctat tggtcttact gacatccact ttgcctttct ctccacaggt gtccactccc
     3301 agttcaatta cagctcttaa ggctagagta ttaatacgac tcactatagg gctagcgatc
     3361 gccatggacc acctcggggc gtccctctgg ccccaggtcg gctccctttg tctcctgctc
     3421 gctggggccg cctgggcgcc cccgcctaac ctcccggacc ccaagttcga gagcaaagcg
     3481 gccttgctgg cggcccgggg gcccgaagag cttctgtgct tcaccgagcg gttggaggac
     3541 ttggtgtgtt tctgggagga agcggcgagc gctggggtgg gcccgggcaa ctacagcttc
     3601 tcctaccagc tcgaggatga gccatggaag ctgtgtcgcc tgcaccaggc tcccacggct
     3661 cgtggtgcgg tgcgcttctg gtgttcgctg cctacagccg acacgtcgag cttcgtgccc
     3721 ctagagttgc gcgtcacagc agcctccggc gctccgcgat atcaccgtgt catccacatc
     3781 aatgaagtag tgctcctaga cgcccccgtg gggctggtgg cgcggttggc tgacgagagc
     3841 ggccacgtag tgttgcgctg gctcccgccg cctgagacac ccatgacgtc tcacatccgc
     3901 tacgaggtgg acgtctcggc cggcaacggc gcagggagcg tacagagggt ggagatcctg
     3961 gagggccgca ccgagtgtgt gctgagcaac ctgcggggcc ggacgcgcta caccttcgcc
     4021 gtccgcgcgc gtatggctga gccgagcttc ggcggcttct ggagcgcctg gtcggagcct
     4081 gtgtcgctgc tgacgcctag cgacctggac cccctcatcc tgacgctctc cctcatcctc
     4141 gtggtcatcc tggtgctgct gaccgtgctc gcgctgctct cccaccgccg ggctctgaag
     4201 cagaagatct ggcctggcat cccgagccca gagagcgagt ttgaaggcct cttcaccacc
     4261 cacaagggta acttccagct gtggctgtac cagaatgatg gctgcctgtg gtggagcccc
     4321 tgcaccccct tcacggagga cccacctgct tccctggaag tcctctcaga gcgctgctgg
     4381 gggacgatgc aggcagtgga gccggggaca gatgatgagg gccccctgct ggagccagtg
     4441 ggcagtgagc atgcccagga tacctatctg gtgctggaca aatggttgct gccccggaac
     4501 ccgcccagtg aggacctccc agggcctggt ggcagtgtgg acatagtggc catggatgaa
     4561 ggctcagaag catcctcctg ctcatctgct ttggcctcga agcccagccc agagggagcc
     4621 tctgctgcca gctttgagta cactatcctg gaccccagct cccagctctt gcgtccatgg
     4681 acactgtgcc ctgagctgcc ccctacccca ccccacctaa agtacctgta ccttgtggta
     4741 tctgactctg gcatctcaac tgactacagc tcaggggact cccagggagc ccaagggggc
     4801 ttatccgatg gcccctactc caacccttat gagaacagcc ttatcccagc cgctgagcct
     4861 ctgcccccca gctatgtggc ttgctctgtt tctcaaggca gttcaggtgg tggcgggagc
     4921 ggaggtggag gctcgagcgg tgtgaccggc taccggctgt tcgaggagat tctgtaaggc
     4981 cgcgactcta gagtcgacct gcaggcatgc aagctgattt ggctgctgcc accgctgagc
     5041 aaggccgctt cgagcagaca tgataagata cattgatgag tttggacaaa ccacaactag
     5101 aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac
     5161 cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt
     5221 tcagggggag atgtgggagg tttttttaag caagtaaaac ctctacaaat gtggtaaaat
     5281 cttaattaag aattctaata taacttcgta tagcatacat tatacgaagt tatggatcca
     5341 ctgttgacaa ttaatcatcg gcatagtata tcggcatagt ataatacgac aaggtgagga
     5401 actaaaccca ggaggcagat catgattgaa caagatggat tgcacgcagg ttctccggcc
     5461 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     5521 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     5581 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     5641 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     5701 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     5761 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     5821 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     5881 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     5941 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     6001 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     6061 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     6121 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     6181 atcgccttct atcgccttct tgacgagttc ttctgattcg aaatgaccga ccaagcgacg
     6241 cccaaccggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     6301 acgcaggaaa gaacatgtca aacatgagaa ttacaactta tatcgtatgg ggctgacttc
     6361 aggtgctaca tttgaagaga taaattgcac tgaaatctag aaatatttta tctgattaat
     6421 aagatgatct tcttgagatc gttttggtct gcgcgtaatc tcttgctctg aaaacgaaaa
     6481 aaccgccttg cagggcggtt tttcgaaggt tctctgagct accaactctt tgaaccgagg
     6541 taactggctt ggaggagcgc agtcaccaaa acttgtcctt tcagtttagc cttaaccggc
     6601 gcatgacttc aagactaact cctctaaatc aattaccagt ggctgctgcc agtggtgctt
     6661 ttgcatgtct ttccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     6721 actgaacggg gggttcgtgc atacagtcca gcttggagcg aactgcctac ccggaactga
     6781 gtgtcaggcg tggaatgaga caaacgcggc cataacagcg gaatgacacc ggtaaaccga
     6841 aaggcaggaa caggagagcg cacgagggag ccgccagggg gaaacgcctg gtatctttat
     6901 agtcctgtcg ggtttcgcca ccactgattt gagcgtcaga tttcgtgatg cttgtcaggg
     6961 gggcggagcc tatggaaaaa cggctttgcc gcggccctct cacttccctg ttaagtatct
     7021 tcctggcatc ttccaggaaa tctccgcccc gttcgtaagc catttccgct cgccgcagtc
     7081 gaacgaccga gcgtagcgag tcagtgagcg aggaagcgga atatatcctg tatcacatat
     7141 tctgctgacg caccggtgca gccttttttc tcctgccaca tgaagcactt cactgacacc
     7201 ctcatcagtg ccaacatagt aagccagtat acactccgct agcgctgatg tccggaatcg
     7261 tcacacaaaa aaccaacaca cagatgtaat gaaaataaag atattttatt gcggccatcg
     7321 tgatggctag agtttagccc tcccacacat aaccagaggg cagcaattca cgaatcccaa
     7381 ctgccgtcgg ctgtccatca ctgtccttca ctatggcttt gatcccagga tgcagatcga
     7441 gaagcacctg tcggcaccgt ccgcaggggc tcaagatgcc cctgttctca tttccgatcg
     7501 cgacgataca agtcaggttg ccagctgccg cagcagcagc agtgcccagc accacgagtt
     7561 ctgcacaagg tcccccagta aaatgatata cattgacacc agtgaagatg cggccgtcgc
     7621 tagagagagc tgcgctggcg acgctgtagt cttcagagat ggggatgctg ttgattgtag
     7681 ccgttgctct ttcaatgagg gtggattctt cttgagacaa aggcttggcc atggtggcgc
     7741 tagtgtcaga agaatcgagc tttttgcaaa agcctaggcc tccaaaaaag cctcctcact
     7801 acttctggaa tagctcagag gccgaggcgg cctcggcctc tgcataaata aaaaaaatta
     7861 gtcagccatg gggcggagaa tgggcggaac tgggcggagt taggggcggg atgggcggag
     7921 ttaggggcgg gactatggtt gctgactaat tgagatgcat gctttgcata cttctgcctg
     7981 ctggggagcc tggggacttt ccacacctgg ttgctgacta attgagatgc atgctttgca
     8041 tacttctgcc tgctggggag cctggggact ttccacaccc taactgacac acattccaca
     8101 gctggttctt tccgcctcag aaggtaccta accaagttcc tctttcagag gttatttcag
     8161 gccatggtgc tgcgc
//
