LOCUS       sequence-481948-        6868 bp    DNA     circular SYN 04-AUG-2025
DEFINITION  Express CRBN-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 6868)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6868)
  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            "Unpublished"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: CRBN-NanoLuc Vector
FEATURES             Location/Qualifiers
     source          1..6868
                     /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"
     CDS             complement(275..787)
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     promoter        complement(2173..2191)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2174..2191)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        2311..2614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2769..2789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        2822..2869
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2901..3560
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2967..2986)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     rep_origin      complement(3732..4320)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3812..3831)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4446..4494
                     /note="synthetic polyadenylation signal"
     CDS             complement(4518..5312)
                     /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(4629..4648)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5239..5258
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(5690..6868)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          5699..6637
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(5722..5742)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
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 ggccttacgc cagaatgcgt tcgcacagcc
      301 gccagccggt cactccgttg atggttactc ggaacagcag ggagccgtcg gggttgatca
      361 ggcgctcgtc gataattttg ttgccgttcc acagggtccc tgttacagtg atctttttgc
      421 cgtcgaacac ggcgatgcct tcatacggcc gtccgaaata gtcgatcatg ttcggcgtaa
      481 ccccgtcgat taccagtgtg ccatagtgca ggatcacctt aaagtgatga tcatccacag
      541 ggtacaccac cttaaaaatt ttttcgatct ggcccatttg gtcgccgctc agaccttcat
      601 acgggatgat gacatggatg tcgatcttca gcccattttc accgctcagg acaatccttt
      661 ggatcggagt tacggacacc ccgagattct gaaacaaact ggacacacct ccctgttcaa
      721 ggacttggtc caggttgtag ccggctgtct gtcgccagtc cccaacgaaa tcttcgagtg
      781 tgaagacgcc gctcgagccg agagaaacca agcaaagtat tactttgtct ggacttattt
      841 catcttcagt gtctgggatc gtgggcaaca gagcagatcg cgttaagccc caaaattttt
      901 gaggtgacat gtcttttttg gtggccgtaa acttccatcc aatatggctt gcacagatct
      961 tacactgggc aacagtccag gcatacccag gaaaccagct gtgttctgta gaaggccggc
     1021 ctatcagatt caagttgcaa gccttataca cagtaagtgt ctcatgcaca tatccatgag
     1081 gattcacata agctgccatc ggcccacata aggataaact gaatatttca tttttggttg
     1141 ttatttctgt ttcttgacat tgtttacagc aaagggaagt acatttattc ataatgtcta
     1201 attcacagcg aagtcgctgg atagcactgc caattttaag gagctgaatt ctcaatacat
     1261 catcaatagg aagacaagca gctactctgt aagaaaaatc tattggattt gaaggaagag
     1321 aatcatcttt tagattttca tcccattcac gtagctgttt cttgattctg tccattaagg
     1381 tctcagcatc atataaggaa tacagccagc gaggccatga agttagattt gcacaatgaa
     1441 actttctctt ctggtatttc tgccaccatt tatatgaaca ttggtcttct cttgagacag
     1501 gttttgaagg aaatatctgg cacttattga gggattctaa ttgaactgca gacatggttg
     1561 aaggcaacac acattcggga agaatttgca ctttagcttg ctggattcca tctgactgtg
     1621 ttcttagctc aaggactttg aacctttgtc ttccaattgc tttcactttc actatctcaa
     1681 ttccaaaatc ctgttcttct cgataggcat atatctctgc tgttgttcca aactgtgctt
     1741 ccctttcctg tacattgctg tatgcaagaa cagcaaaggt tctatctttc tgaattaaat
     1801 tccgcaccat actgacttct tgagggtgaa aaagctgaag aggtaatgtc tgtccgggaa
     1861 tcaggatcat catcacttgt ggaagaactg gaatcacctg acagctgtcg tcatcgtgca
     1921 aagtcctgcc atgaaattct tccatatcag cacctaggta tgtatgtgat gtcggcagac
     1981 tggtgtcaaa atttatgatg tttggttttt tggcttcttt actatcctgg tcttcaactt
     2041 ccatttcatc ttcttcctca ctctctgcag gcaggagcgg caggtggttg cccatgttgt
     2101 gcgcagcgtc ctgctgatct ccttcgccgg ccatggcgat cgctagctgc aaaaagaaca
     2161 agtagcttgt attctatagt gtcacctaaa tgggcgatct gacggttcac taaaccagct
     2221 ctgcttatat aggcctccct ggcgcgccac tagttattaa tagtaatcaa ttacggggtc
     2281 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
     2341 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
     2401 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
     2461 cttggcagta catcaagtgt atcatatgcc aagtccgccc cctattgacg tcaatgacgg
     2521 taaatggccc gcctggcatt atgcccagta catgacctta cgggactttc ctacttggca
     2581 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacaccaa
     2641 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
     2701 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta ataaccccgc
     2761 cccgttgacg caaatgggcg gtaggcgtgt acggtgggag gtcaaatcga attctaatac
     2821 tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa
     2881 ctaaacccag gaggcagatc atggagaaaa aaatcactgg atataccacc gttgatatat
     2941 cccaatggca tcgtaaagaa cattttgagg catttcagtc agttgctcaa tgtacctata
     3001 accagaccgt tcagctggat attacggcct ttttaaagac cgtaaagaaa aataagcaca
     3061 agttttatcc ggcctttatt cacattcttg cccgcctgat gaatgctcat ccggaattcc
     3121 gtatggcaat gaaagacggt gagctggtga tatgggatag tgttcaccct tgttacaccg
     3181 ttttccatga gcaaactgaa acgttttcat cgctctggag tgaataccac gacgatttcc
     3241 ggcagtttct acacatatat tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt
     3301 tccctaaagg gtttattgag aatatgtttt tcgtctcagc caatccctgg gtgagtttca
     3361 ccagttttga tttaaacgtg gccaatatgg acaacttctt cgcccccgtt ttcactatgg
     3421 gcaaatatta tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag gttcatcatg
     3481 ccgtctgtga tggcttccat gtcggcagaa tgcttaatga attacaacag tactgcgatg
     3541 agtggcaggg cggggcgtaa gcgggactct ggggttcgaa atgaccgacc aagcgacgcc
     3601 caaccggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     3661 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     3721 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     3781 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     3841 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     3901 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     3961 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4021 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4081 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4141 aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg cgctctgctg
     4201 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4261 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatttcaa
     4321 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     4381 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt atagtccgga
     4441 atcgtcacac aaaaaaccaa cacacagatg taatgaaaat aaagatattt tattgcggcc
     4501 atcgtgatgg ctagagttca gaagaactcg tcaagaaggc gatagaaggc gatgcgctgc
     4561 gaatcgggag cggcgatacc gtaaagcacg aggaagcggt cagcccattc gccgccaagc
     4621 tcttcagcaa tatcacgggt agccaacgct atgtcctgat agcggtccgc cacacccagc
     4681 cggccacagt cgatgaatcc agaaaagcgg ccattttcca ccatgatatt cggcaagcag
     4741 gcatcgccgt gggtcacgac gagatcctcg ccgtcgggca tgcgcgcctt gagcctggcg
     4801 aacagttcgg ctggcgcgag cccctgatgc tcttcgtcca gatcatcctg atcgacaaga
     4861 ccggcttcca tccgagtacg tgctcgctcg atgcgatgtt tcgcttggtg gtcgaatggg
     4921 caggtagccg gatcaagcgt atgcagccgc cgcattgcat cagccatgat ggatactttc
     4981 tcggcaggag caaggtgaga tgacaggaga tcctgccccg gcacttcgcc caatagcagc
     5041 cagtcccttc ccgcttcagt gacaacgtcg agcacagctg cgcaaggaac gcccgtcgtg
     5101 gccagccacg atagccgcgc tgcctcgtcc tgcagttcat tcagggcacc ggacaggtcg
     5161 gtcttgacaa aaagaaccgg gcgcccctgc gctgacagcc ggaacacggc ggcatcagag
     5221 cagccgattg tctgttgtgc ccagtcatag ccgaatagcc tctccaccca agcggccgga
     5281 gaacctgcgt gcaatccatc ttgttcaatc atggtggcat cgattcccgg tcaacatcca
     5341 gtacacctcc ggcacaccgg cttccccaag ggcgccacgc tctcccacca caacatcctc
     5401 aacaacggct actgggtggg ccgcacggtc ggctacacgg agcaggaccg ggtgtgtctg
     5461 ccggtgccct tctaccactg cttcggcatg gtcatgggca acttgggcgc cacctcccac
     5521 gccgcctgca tcgtcatccc ggccccgtcc ttcgagccgg cggcgacgct ggaggcggtg
     5581 cagcgggaga ggtgcacgtc cctgtacggc gtccccacga agccgaattc caccacactg
     5641 gactagatgc atgctcgaca gtgtgatgga tatctgcaga attcggcttt cacgacacct
     5701 gaaatggaag aaaaaaactt tgaaccactg tctgaggctt gagaatgaac caagatccaa
     5761 actcaaaaag ggcaaattcc aaggagaatt acatcaagtg ccaagctggc ctaacttcag
     5821 tctccaccca ctcagtgtgg ggaaactcca tcgcataaaa cccctccccc caacctaaag
     5881 acgacgtact ccaaaagctc gagaactaat cgaggtgcct ggacggcgcc cggtactccg
     5941 tggagtcaca tgaagcgacg gctgaggacg gaaaggccct tttcctttgt gtgggtgact
     6001 cacccgcccg ctctcccgag cgccgcgtcc tccattttga gctgcctgca gcagggccgg
     6061 gaagcggcca tctttccgct cacgcaactg gtgccgaccg ggccagcctt gccgcccagg
     6121 gcggggcgat acacggcggc gcgaggccag gcaccagagc aggccggcca gcttgagact
     6181 acccccgtcc gattctcggt ggccgcgctc gcaggccccg cctcgccgaa catgtgcgct
     6241 gggacgcacg ggccccgtcg ccgcccgcgg ccccaaaaac cgaaatacca gtgtgcagat
     6301 cttggcccgc atttacaaga ctatcttgcc agaaaaaaag cgtcgcagca ggtcatcaaa
     6361 aattttaaat ggctagagac ttatcgaaag cagcgagaca ggcgcgaagg tgccaccaga
     6421 ttcgcacgcg gcggccccag cgcccaggcc aggcctcaac tcaagcacga ggcgaagggg
     6481 ctccttaagc gcaaggcctc gaactctccc acccacttcc aacccgaagc tcgggatcaa
     6541 gaatcacgta ctgcagccag gtggaagtaa ttcaaggcac gcaagggcca taacccgtaa
     6601 agaggccagg cccgcgggaa ccacacacgg cacttacctg tgttctggcg gcaaacccgt
     6661 tgcgaaaaag aacgttcacg gcgactactg cacttatata cggttctccc ccaccctcgg
     6721 gaaaaaggcg gagccagtac acgacatcac tttcccagtt taccccgcgc caccttctct
     6781 aggcaccggt tcaattgccg acccctcccc ccaacttctc ggggactgtg ggcgatgtgc
     6841 gctctgccca ctgacgggca ccggagcc
//
