LOCUS       sequence-476609-        5462 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express CHMP4B-LgBiT 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 5462)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5462)
  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            "Unpublished"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: CHMP4B-LgBiT Fusion Vector
FEATURES             Location/Qualifiers
     source          1..5462
                     /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"
     promoter        complement(1516..1534)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1517..1534)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        1654..1957
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2112..2132
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        2165..2212
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2244..3038
                     /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(2298..2317)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2908..2927
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(3196..3784)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3276..3295)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3910..3958
                     /note="synthetic polyadenylation signal"
     CDS             complement(3982..5019)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             3995..4003
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     promoter        complement(5050..5407)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5064..5199
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        5216..5407
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5364..5384
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
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 ctagagtcgc ggccttagct gttgatggtt actcggaaca
      301 gcatggagcc gtcgggggtg atcaggcgct cgtcgataat tttgttgccg ttccacaggg
      361 tccctgttac agtgatcttt ttgccgtcga acacggcgat gccttcatac ggccgtccga
      421 aatagttcag catgttcggc gtaaccccgt cgattaccag tgtgccatag ggcaggatca
      481 ccttaaagtg atgatcatcc acagggtaca ccaccttaaa cacctcttcg atctgggcca
      541 tttggtcggc gctcagacct tcatacggga tgatgacatg gatgtcgatc ttcagggcat
      601 tttcaccgct ccggacaatc ctttggatcg gagttacgga cacggcgaga ttctgcagca
      661 aactggacac acctccctgt tcaaggactt ggtccaggtt gtaggcggct gtctgttccc
      721 agtccccaac gaaatcttcg agtgtgaaga caccgctcga gcctccacct ccgctcccgc
      781 caccacctga actgccttga gaaaccatgg atccagccca gttctccaat tccttcatgt
      841 cgtcgtcctc ctcttctttc ttcttggcgg gttttgatgg tagggctata gagggaacat
      901 ttggtagagg gactgtttcg ggtccactga tttccagcaa attcttgtct agttcctcct
      961 gttctagttc ttctaattcc gccatgagct catcctcgtc aaactcttct ccaaacccta
     1021 caggtttcga aattgctgtt gaaatctcct ctgcaagttc ttgctggtca gcaatgtcct
     1081 gcattaactc atcaacttta tcgatgtcca tgttgtcatg ggccgccttc atggccttgg
     1141 cggcatagcc catgttcttg agcacctcgg tgttggtgtt ggcattctcc agggcctccc
     1201 gctggaactc gatggttgat aatgtgccgt cgatctgcgc cagctgcttc tcatacctct
     1261 tcttacgctt cagtgcctgg agggccgcgc gcttgttttt ggtgccgtgc ttcttggcgg
     1321 ccgtcagctc ctgctcgatt ttcttctcca ggaactcctg tttcttgctt aacatctctt
     1381 ccgtgtcccg cagccgctgg atggcctcct ggggggtcgg gccgcccttg ccggccttac
     1441 cccctccagc cccgaacagc ttcccgaaca ccgacatggc gatcgctagc tgcaaaaaga
     1501 acaagtagct tgtattctat agtgtcacct aaatgggcga tctgacggtt cactaaacca
     1561 gctctgctta tataggcctc cctggcgcgc cactagttat taatagtaat caattacggg
     1621 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
     1681 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
     1741 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
     1801 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
     1861 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
     1921 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
     1981 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
     2041 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
     2101 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtcaaat cgaattctaa
     2161 tactgttgac aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag
     2221 gaactaaacc caggaggcag atcatgattg aacaagatgg attgcacgca ggttctccgg
     2281 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     2341 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     2401 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     2461 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     2521 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     2581 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     2641 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     2701 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     2761 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     2821 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     2881 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     2941 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     3001 gcatcgcctt ctatcgcctt cttgacgagt tcttctgatt cgaaatgacc gaccaagcga
     3061 cgcccaaccg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     3121 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     3181 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     3241 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     3301 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     3361 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     3421 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     3481 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     3541 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     3601 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     3661 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     3721 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatt
     3781 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     3841 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttatagtc
     3901 cggaatcgtc acacaaaaaa ccaacacaca gatgtaatga aaataaagat attttattgc
     3961 ggccatcgtg atggctagag tctaaactcg acctacctcc ttagcgcgcg gtcgtgtact
     4021 gggccggcgg ttgccgctgt cggccagcac ctcgacgcag ccgtcggtcc atacggctgc
     4081 gctccggcga gcgatttgtg tgcgaccgac ggtgccggcc ccgctgcgga cgatggcatc
     4141 gcagcggcct tgagcccagg cagcatcgtc gaagttgccg tccacgaggc tctggtagag
     4201 ctgatccagg ccgatgcgca gcatgtaggc tcgcagacga gggctgccgg ccagctcggg
     4261 atgccggcgc tcgaagtagc gagtctgctg ctccatgcaa gccagccagg gccgccagaa
     4321 gaagatgttg gccacctcgt actgactgtc cccgaacata gcttcggacc agtcgattac
     4381 ggcggtgatg cggccgttgt cggtcaggac gttgttgctg ccgaagtcgg catggacgag
     4441 gtggcgcacc tcgggacagt cttcggccca cagcatcagt tcgtccaggg cttgagctac
     4501 gctggcggac acggtgtcgt ccatcacggt ctgccagtgg tagacatggg gatcagcaat
     4561 ggcgcaaatg aaatcccgcc aggttgtgta ctggccgatg ccttggggcc cgaatgggcc
     4621 gaagccgctg gtttgactga ggtcggcggc ggcaatagca tccatagctt cggcgacagg
     4681 ctgtaacaca gcaggcagct ctgtttcggg gaggtcttgg agagtaacgc cttgggcgcg
     4741 tctactgatg cagtatgtca ggctctcgct gaactcgccg atgtccaaca cttcggggat
     4801 gggtagtgca gcgctggcga agtggcggta cacgtagcgg tctttgtaga agccatcagc
     4861 gcagctattc acccgcagta catagccgcg tccgccgaca tcgaagctga aggctcggct
     4921 ctcttcgccc tccgacaact gcatcaggtc gctcacactg tcgaacttct cgatgagaaa
     4981 tttttcaacg ctggtagcgg tgagttcggg cttcttcatg gtggcgctag tgtcagaaga
     5041 atcgagcttt ttgcaaaagc ctaggcctcc aaaaaagcct cctcactact tctggaatag
     5101 ctcagaggcc gaggcggcct cggcctctgc ataaataaaa aaaattagtc agccatgggg
     5161 cggagaatgg gcggaactgg gcggagttag gggcgggatg ggcggagtta ggggcgggac
     5221 tatggttgct gactaattga gatgcatgct ttgcatactt ctgcctgctg gggagcctgg
     5281 ggactttcca cacctggttg ctgactaatt gagatgcatg ctttgcatac ttctgcctgc
     5341 tggggagcct ggggactttc cacaccctaa ctgacacaca ttccacagct ggttctttcc
     5401 gcctcagaag gtacctaacc aagttcctct ttcagaggtt atttcaggcc atggtgctgc
     5461 gc
//
