LOCUS       sequence-473310-        6822 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express RNF43-SmBiT 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 6822)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6822)
  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            "Unpublished"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: RNF43-SmBiT, BiBiT-Ready Vector
FEATURES             Location/Qualifiers
     source          1..6822
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    24..105
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     enhancer        338..641
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        642..845
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     796..816
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          981..1113
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1096..1115
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1157..1176
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1157..1175
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    3711..3832
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3722..3741)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(3947..3980)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        3990..4037
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4069..4863
                     /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(4123..4142)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4733..4752
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      5080..5625
                     /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     5526..5545
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5742..5759
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    5909..5957
                     /note="synthetic polyadenylation signal"
     CDS             complement(5981..6379)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     regulatory      6376..6385
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(6410..6767)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6424..6559
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        6576..6767
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     6724..6744
                     /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 ggattatggc gcgccactag ttattaatag taatcaatta
      301 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
      361 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
      421 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
      481 ctgcccactt ggcagtacat caagtgtatc atatgccaag tccgccccct attgacgtca
      541 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttacgg gactttccta
      601 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
      661 acaccaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
      721 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaata
      781 accccgcccc gttgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
      841 gagctggttt agtgaaccgt cagatcacta gaagctttat tgcggtagtt tatcacagtt
      901 aaattgctaa cgcagtcagt gcttctgaca caacagtctc gaacttaagc tgcagaagtt
      961 ggtcgtgagg cactgggcag gtaagtatca aggttacaag acaggtttaa ggagaccaat
     1021 agaaactggg cttgtcgaga cagagaagac tcttgcgttt ctgataggca cctattggtc
     1081 ttactgacat ccactttgcc tttctctcca caggtgtcca ctcccagttc aattacagct
     1141 cttaaggcta gagtattaat acgactcact atagggctag cgatcgccat gagtggtggc
     1201 caccagctgc agctggctgc cctctggccc tggctgctga tggctaccct gcaggcaggc
     1261 tttggacgca caggactggt actggcagca gcggtggagt ctgaaagatc agcagaacag
     1321 aaagctatta tcagagtgat ccccttgaaa atggacccca caggaaaact gaatctcact
     1381 ttggaaggtg tgtttgctgg tgttgctgaa ataactccag cagaaggaaa attaatgcag
     1441 tcccacccgc tgtacctgtg caatgccagt gatgacgaca atctggagcc tggattcatc
     1501 agcatcgtca agctggagag tcctcgacgg gccccccgcc cctgcctgtc actggctagc
     1561 aaggctcgga tggcgggtga gcgaggagcc agtgctgtcc tctttgacat cactgaggat
     1621 cgagctgctg ctgagcagct gcagcagccg ctggggctga cctggccagt ggtgttgatc
     1681 tggggtaatg acgctgagaa gctgatggag tttgtgtaca agaaccaaaa ggcccatgtg
     1741 aggattgagc tgaaggagcc cccggcctgg ccagattatg atgtgtggat cctaatgaca
     1801 gtggtgggca ccatctttgt gatcatcctg gcttcggtgc tgcgcatccg gtgccgcccc
     1861 cgccacagca ggccggatcc gcttcagcag agaacagcct gggccatcag ccagctggcc
     1921 accaggaggt accaggccag ctgcaggcag gcccggggtg agtggccaga ctcagggagc
     1981 agctgcagct cagcccctgt gtgtgccatc tgtctggagg agttctctga ggggcaggag
     2041 ctacgggtca tttcctgcct ccatgagttc catcgtaact gtgtggaccc ctggttacat
     2101 cagcatcgga cttgccccct ctgcatgttc aacatcacag agggagattc attttcccag
     2161 tccctgggac cctctcgatc ttaccaagaa ccaggtcgaa gactccacct cattcgccag
     2221 catcccggcc atgcccacta ccacctccct gctgcctacc tgttgggccc ttcccggagt
     2281 gcagtggctc ggcccccacg acctggtccc ttcctgccat cccaggagcc aggcatgggc
     2341 cctcggcatc accgcttccc cagagctgca catccccggg ctccaggaga gcagcagcgc
     2401 ctggcaggag cccagcaccc ctatgcacaa ggctggggac tgagccacct ccaatccacc
     2461 tcacagcacc ctgctgcttg cccagtgccc ctacgccggg ccaggccccc tgacagcagt
     2521 ggatctggag aaagctattg cacagaacgc agtgggtacc tggcagatgg gccagccagt
     2581 gactccagct cagggccctg tcatggctct tccagtgact ctgtggtcaa ctgcacggac
     2641 atcagcctac agggggtcca tggcagcagt tctactttct gcagctccct aagcagtgac
     2701 tttgaccccc tagtgtactg cagccctaaa ggggatcccc agcgagtgga catgcagcct
     2761 agtgtgacct ctcggcctcg ttccttggac tcggtggtgc ccacagggga aacccaggtt
     2821 tccagccatg tccactacca ccgccaccgg caccaccact acaaaaagcg gttccagtgg
     2881 catggcagga agcctggccc agaaaccgga gtcccccagt ccaggcctcc tattcctcgg
     2941 acacagcccc agccagagcc accttctcct gatcagcaag tcaccagatc caactcagca
     3001 gccccttcgg ggcggctctc taacccacag tgccccaggg ccctccctga gccagcccct
     3061 ggcccagttg acgcctccag catctgcccc agtaccagca gtctgttcaa cttgcaaaaa
     3121 tccagcctct ctgcccgaca cccacagagg aaaaggcggg ggggtccctc cgagcccacc
     3181 cctggctctc ggccccagga tgcaactgtg cacccagctt gccagatttt tccccattac
     3241 acccccagtg tggcatatcc ttggtcccca gaggcacacc ccttgatctg tggacctcca
     3301 ggcctggaca agaggctgct accagaaacc ccaggcccct gttactcaaa ttcacagcca
     3361 gtgtggttgt gcctgactcc tcgccagccc ctggaaccac atccacctgg ggaggggcct
     3421 tctgaatgga gttctgacac cgcagagggc aggccatgcc cttatccgca ctgccaggtg
     3481 ctgtcggccc agcctggctc agaggaggaa ctcgaggagc tgtgtgaaca ggctgtttct
     3541 caaggcagtt caggtggtgg cgggagcgga ggtggaggct cgagcggtgt gaccggctac
     3601 cggctgttcg aggagattct gtaaggccgc gactctagag tcgacctgca ggcatgcaag
     3661 ctgatttggc tgctgccacc gctgagcaag gccgcttcga gcagacatga taagatacat
     3721 tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat
     3781 ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa
     3841 caattgcatt cattttatgt ttcaggttca gggggagatg tgggaggttt ttttaagcaa
     3901 gtaaaacctc tacaaatgtg gtaaaatctt aattaagaat tctaatataa cttcgtatag
     3961 catacattat acgaagttat ggatccactg ttgacaatta atcatcggca tagtatatcg
     4021 gcatagtata atacgacaag gtgaggaact aaacccagga ggcagatcat gattgaacaa
     4081 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     4141 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     4201 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     4261 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     4321 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     4381 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     4441 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     4501 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     4561 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     4621 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     4681 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     4741 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     4801 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     4861 tgattcgaaa tgaccgacca agcgacgccc aaccggtatc agctcactca aaggcggtaa
     4921 tacggttatc cacagaatca ggggataacg caggaaagaa catgtcaaac atgagaatta
     4981 caacttatat cgtatggggc tgacttcagg tgctacattt gaagagataa attgcactga
     5041 aatctagaaa tattttatct gattaataag atgatcttct tgagatcgtt ttggtctgcg
     5101 cgtaatctct tgctctgaaa acgaaaaaac cgccttgcag ggcggttttt cgaaggttct
     5161 ctgagctacc aactctttga accgaggtaa ctggcttgga ggagcgcagt caccaaaact
     5221 tgtcctttca gtttagcctt aaccggcgca tgacttcaag actaactcct ctaaatcaat
     5281 taccagtggc tgctgccagt ggtgcttttg catgtctttc cgggttggac tcaagacgat
     5341 agttaccgga taaggcgcag cggtcggact gaacgggggg ttcgtgcata cagtccagct
     5401 tggagcgaac tgcctacccg gaactgagtg tcaggcgtgg aatgagacaa acgcggccat
     5461 aacagcggaa tgacaccggt aaaccgaaag gcaggaacag gagagcgcac gagggagccg
     5521 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacca ctgatttgag
     5581 cgtcagattt cgtgatgctt gtcagggggg cggagcctat ggaaaaacgg ctttgccgcg
     5641 gccctctcac ttccctgtta agtatcttcc tggcatcttc caggaaatct ccgccccgtt
     5701 cgtaagccat ttccgctcgc cgcagtcgaa cgaccgagcg tagcgagtca gtgagcgagg
     5761 aagcggaata tatcctgtat cacatattct gctgacgcac cggtgcagcc ttttttctcc
     5821 tgccacatga agcacttcac tgacaccctc atcagtgcca acatagtaag ccagtataca
     5881 ctccgctagc gctgatgtcc ggaatcgtca cacaaaaaac caacacacag atgtaatgaa
     5941 aataaagata ttttattgcg gccatcgtga tggctagagt ttagccctcc cacacataac
     6001 cagagggcag caattcacga atcccaactg ccgtcggctg tccatcactg tccttcacta
     6061 tggctttgat cccaggatgc agatcgagaa gcacctgtcg gcaccgtccg caggggctca
     6121 agatgcccct gttctcattt ccgatcgcga cgatacaagt caggttgcca gctgccgcag
     6181 cagcagcagt gcccagcacc acgagttctg cacaaggtcc cccagtaaaa tgatatacat
     6241 tgacaccagt gaagatgcgg ccgtcgctag agagagctgc gctggcgacg ctgtagtctt
     6301 cagagatggg gatgctgttg attgtagccg ttgctctttc aatgagggtg gattcttctt
     6361 gagacaaagg cttggccatg gtggcgctag tgtcagaaga atcgagcttt ttgcaaaagc
     6421 ctaggcctcc aaaaaagcct cctcactact tctggaatag ctcagaggcc gaggcggcct
     6481 cggcctctgc ataaataaaa aaaattagtc agccatgggg cggagaatgg gcggaactgg
     6541 gcggagttag gggcgggatg ggcggagtta ggggcgggac tatggttgct gactaattga
     6601 gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca cacctggttg
     6661 ctgactaatt gagatgcatg ctttgcatac ttctgcctgc tggggagcct ggggactttc
     6721 cacaccctaa ctgacacaca ttccacagct ggttctttcc gcctcagaag gtacctaacc
     6781 aagttcctct ttcagaggtt atttcaggcc atggtgctgc gc
//
