LOCUS       sequence-479848-        6046 bp    DNA     circular SYN 04-AUG-2025
DEFINITION  Express LgBiT-CCNE1 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 6046)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6046)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 4, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: LgBiT-CCNE1 Fusion Vector
FEATURES             Location/Qualifiers
     source          1..6046
                     /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(2100..2118)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2101..2118)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        2238..2541
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2696..2716
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        2749..2796
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2828..3622
                     /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(2882..2901)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3492..3511
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(3780..4368)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3860..3879)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4494..4542
                     /note="synthetic polyadenylation signal"
     CDS             complement(4566..5603)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4579..4587
                     /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(5634..5991)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5648..5783
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        5800..5991
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5948..5968
                     /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 ctagaggatc cccgggtacc gagcccgaat tcgtttaaac
      301 cgccatttcc ggcccgctgc tctgcttctt accgctctgt ggcggggtga ggagcccact
      361 ggggagagga gaagccctat tttgttcaga caacatggct ttctttgctc gggctttgtc
      421 cagcaaatcc aagctgtctc tgtgggtctg tatgttgtgt gcatcttcat cagcgacgcc
      481 cctgaagtgc ttcagttttg agctccccgt ctcccttata accatggcaa atggaaccat
      541 ccacttgaca cagttctcta tgtcgcacca ctgataccct gaaacctttt gcatcaattc
      601 agatgacgag aaatgataca aggccgaagc agcaagtata ccataaggaa attcaaggca
      661 gtcaacatcc aggacacaga gatccaacag ctctgcaatc tgtataaaga tttgctgggg
      721 atactgcggc agtagcactt catgtaagtc atttagatat gcaacctgca tgtatacatt
      781 cagccaggac acaatagtca ggggacttaa acgccactta agggccttca taatcattaa
      841 ttccatggtg agaatttcat ctcctgaaca agctccatct gtcacatacg caaactggtg
      901 caactttgga ggatagattt cctcaagttt ggctgcaata aataaagatg aaatcccaat
      961 aagctgtaaa agagttttta caacattttc ttgtgtcgcc atataccggt caaagaaatc
     1021 ttgtgccaag taaaaggtct ccctgtgaag tttatagact tcacacacct ccattaacca
     1081 atccagaaga attgctcgca tttttggctg cagaagaggg tgttgctcaa gaaagtgctg
     1141 atcccttaag tatgtctttt ccttgtttaa catgattttc cagacttcct ctctatttgc
     1201 ccagctcagt acaggcagcg gggagcctct ggatggtgca ataatccgag gcttgcacgt
     1261 tgagtttggg taaacccggt catcatcttc tttgtcaggt gtggggatca gggagcaggg
     1321 gtctgcacag actgcattat tgtcccaagg ctggctccca cactggtccc tcgccgtcct
     1381 gtcgattttg gccatttctt catctggatc ctgcaaaaaa acggtcacgt ttgccttcct
     1441 cttcctggag cgagccgaga actccgcgcc gccgtcctcc ttcatggtgt cccgctcctt
     1501 cgcatcccgc tccctgcgct ccctcggcat ggcgatcgca ccgctcgagc ctccacctcc
     1561 gctcccgcca ccaccggagc tcccactgtt gatggttact cggaacagca tggagccgtc
     1621 gggggtgatc aggcgctcgt cgataatttt gttgccgttc cacagggtcc ctgttacagt
     1681 gatctttttg ccgtcgaaca cggcgatgcc ttcatacggc cgtccgaaat agttcagcat
     1741 gttcggcgta accccgtcga ttaccagtgt gccatagggc aggatcacct taaagtgatg
     1801 atcatccaca gggtacacca ccttaaacac ctcttcgatc tgggccattt ggtcggcgct
     1861 cagaccttca tacgggatga tgacatggat gtcgatcttc agggcatttt caccgctccg
     1921 gacaatcctt tggatcggag ttacggacac ggcgagattc tgcagcaaac tggacacacc
     1981 tccctgttca aggacttggt ccaggttgta ggcggctgtc tgttcccagt ccccaacgaa
     2041 atcttcgagt gtgaagacca tggtgagcgc tagctgcaaa aagaacaagt agcttgtatt
     2101 ctatagtgtc acctaaatgg gcgatctgac ggttcactaa accagctctg cttatatagg
     2161 cctccctggc gcgccactag ttattaatag taatcaatta cggggtcatt agttcatagc
     2221 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
     2281 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
     2341 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
     2401 caagtgtatc atatgccaag tccgccccct attgacgtca atgacggtaa atggcccgcc
     2461 tggcattatg cccagtacat gaccttacgg gactttccta cttggcagta catctacgta
     2521 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acaccaatgg gcgtggatag
     2581 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
     2641 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaata accccgcccc gttgacgcaa
     2701 atgggcggta ggcgtgtacg gtgggaggtc aaatcgaatt ctaatactgt tgacaattaa
     2761 tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta aacccaggag
     2821 gcagatcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     2881 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     2941 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     3001 tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     3061 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     3121 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     3181 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     3241 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     3301 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat
     3361 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     3421 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     3481 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     3541 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     3601 ccttcttgac gagttcttct gattcgaaat gaccgaccaa gcgacgccca accggtatca
     3661 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     3721 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     3781 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     3841 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     3901 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     3961 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     4021 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     4081 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     4141 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     4201 aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc
     4261 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     4321 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatttcaaga agatcctttg
     4381 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     4441 atgagattat caaaaaggat cttcacctag atccttttat agtccggaat cgtcacacaa
     4501 aaaaccaaca cacagatgta atgaaaataa agatatttta ttgcggccat cgtgatggct
     4561 agagtctaaa ctcgacctac ctccttagcg cgcggtcgtg tactgggccg gcggttgccg
     4621 ctgtcggcca gcacctcgac gcagccgtcg gtccatacgg ctgcgctccg gcgagcgatt
     4681 tgtgtgcgac cgacggtgcc ggccccgctg cggacgatgg catcgcagcg gccttgagcc
     4741 caggcagcat cgtcgaagtt gccgtccacg aggctctggt agagctgatc caggccgatg
     4801 cgcagcatgt aggctcgcag acgagggctg ccggccagct cgggatgccg gcgctcgaag
     4861 tagcgagtct gctgctccat gcaagccagc cagggccgcc agaagaagat gttggccacc
     4921 tcgtactgac tgtccccgaa catagcttcg gaccagtcga ttacggcggt gatgcggccg
     4981 ttgtcggtca ggacgttgtt gctgccgaag tcggcatgga cgaggtggcg cacctcggga
     5041 cagtcttcgg cccacagcat cagttcgtcc agggcttgag ctacgctggc ggacacggtg
     5101 tcgtccatca cggtctgcca gtggtagaca tggggatcag caatggcgca aatgaaatcc
     5161 cgccaggttg tgtactggcc gatgccttgg ggcccgaatg ggccgaagcc gctggtttga
     5221 ctgaggtcgg cggcggcaat agcatccata gcttcggcga caggctgtaa cacagcaggc
     5281 agctctgttt cggggaggtc ttggagagta acgccttggg cgcgtctact gatgcagtat
     5341 gtcaggctct cgctgaactc gccgatgtcc aacacttcgg ggatgggtag tgcagcgctg
     5401 gcgaagtggc ggtacacgta gcggtctttg tagaagccat cagcgcagct attcacccgc
     5461 agtacatagc cgcgtccgcc gacatcgaag ctgaaggctc ggctctcttc gccctccgac
     5521 aactgcatca ggtcgctcac actgtcgaac ttctcgatga gaaatttttc aacgctggta
     5581 gcggtgagtt cgggcttctt catggtggcg ctagtgtcag aagaatcgag ctttttgcaa
     5641 aagcctaggc ctccaaaaaa gcctcctcac tacttctgga atagctcaga ggccgaggcg
     5701 gcctcggcct ctgcataaat aaaaaaaatt agtcagccat ggggcggaga atgggcggaa
     5761 ctgggcggag ttaggggcgg gatgggcgga gttaggggcg ggactatggt tgctgactaa
     5821 ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt tccacacctg
     5881 gttgctgact aattgagatg catgctttgc atacttctgc ctgctgggga gcctggggac
     5941 tttccacacc ctaactgaca cacattccac agctggttct ttccgcctca gaaggtacct
     6001 aaccaagttc ctctttcaga ggttatttca ggccatggtg ctgcgc
//
