LOCUS       sequence-479806-        6477 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Express CRBN-HaloTag 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 6477)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6477)
  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            "Unpublished"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: NanoBRET Assay Vector, CRBN-HaloTag
FEATURES             Location/Qualifiers
     source          1..6477
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    828..876
                     /note="synthetic polyadenylation signal"
     CDS             complement(940..1734)
                     /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(1051..1070)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1661..1680
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1766..1813)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(1828..2185)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1842..1977
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        1994..2185
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     2142..2162
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    2430..2551
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2467..2486)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2521..2540
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2588..2606
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(2694..3578)
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     CDS             complement(3594..3614)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     primer_bind     complement(4974..4993)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4975..4993)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5035..5054)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          5037..5169
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     promoter        complement(5305..5508)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(5334..5354)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5509..5888
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     misc_feature    6079..6362
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
ORIGIN
        1 ttgaaatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgggta
      721 gaaagccagt ccgcagaaac ggtgctgacc ccggatgaat gtcagctact gggctatctg
      781 gacaagggaa aacgcaagcg caaagaggat ccttatcgct atcgattcac acaaaaaacc
      841 aacacacaga tgtaatgaaa ataaagatat tttattgcgg ccatcgtgat ggcaggttgg
      901 gcgtcgcttg gtcggtcatt tcgaacccca gagtcccgct cagaagaact cgtcaagaag
      961 gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca cgaggaagcg
     1021 gtcagcccat tcgccgccaa gctcttcagc aatatcacgg gtagccaacg ctatgtcctg
     1081 atagcggtcc gccacaccca gccggccaca gtcgatgaat ccagaaaagc ggccattttc
     1141 caccatgata ttcggcaagc aggcatcgcc atgggtcacg acgagatcct cgccgtcggg
     1201 catgcgcgcc ttgagcctgg cgaacagttc ggctggcgcg agcccctgat gctcttcgtc
     1261 cagatcatcc tgatcgacaa gaccggcttc catccgagta cgtgctcgct cgatgcgatg
     1321 tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc gccgcattgc
     1381 atcagccatg atggatactt tctcggcagg agcaaggtga gatgacagga gatcctgccc
     1441 cggcacttcg cccaatagca gccagtccct tcccgcttca gtgacaacgt cgagcacagc
     1501 tgcgcaagga acgcccgtcg tggccagcca cgatagccgc gctgcctcgt cctgcagttc
     1561 attcagggca ccggacaggt cggtcttgac aaaaagaacc gggcgcccct gcgctgacag
     1621 ccggaacacg gcggcatcag agcagccgat tgtctgttgt gcccagtcat agccgaatag
     1681 cctctccacc caagcggccg gagaacctgc gtgcaatcca tcttgttcaa tcatgatctg
     1741 cctcctgggt ttagttcctc accttgtcgt attatactat gccgatatac tatgccgatg
     1801 attaattgtc aacagttaat taagcttttt gcaaaagcct aggcctccaa aaaagcctcc
     1861 tcactacttc tggaatagct cagaggccga ggcggcctcg gcctctgcat aaataaaaaa
     1921 aattagtcag ccatggggcg gagaatgggc ggaactgggc ggagttaggg gcgggatggg
     1981 cggagttagg ggcgggacta tggttgctga ctaattgaga tgcatgcttt gcatacttct
     2041 gcctgctggg gagcctgggg actttccaca cctggttgct gactaattga gatgcatgct
     2101 ttgcatactt ctgcctgctg gggagcctgg ggactttcca caccctaact gacacacatt
     2161 ccacagctgg ttctttccgc ctcagaaggt acctaaccaa gttcctcttt cagaggttat
     2221 ttcaggccat ggtgctgcgc agatccgcgt atgcggtgtg aaataccgca cagatgcgta
     2281 aggagaaaat accgcatcag gaaattgtaa gcgttaatat tttgttaaaa ttcgatttta
     2341 ccacatttgt agaggtttta cttgcttaaa aaaacctccc acatctcccc ctgaacctga
     2401 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
     2461 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     2521 gtggtttgtc caaactcatc aatgtatctt atcatgtctg ctcgaagcgg ccgccccaag
     2581 gggttatgct agttattgct cagcggtggc agcagccaac tcagcttcct ttcgggcttt
     2641 gttagcagcc ggatcagctt gcatgcctgc aggtcgactc tagaattcta ttaaccggaa
     2701 atctccagag tagacagcca gcgcgcgatc tcgctgccga tcaggtccgg gttgtcttct
     2761 tgcagcagat tcagacccgg gccgatgtcc acagccttgc agttaggcag gcttttggcc
     2821 aggcgagcgg cttcggccgg tgggatcaga acgcctgggg tgccccagaa cagcagcttc
     2881 gggacagggg actggtgcag ccagtccatg tattcttcga ccagcgcgac gatgttcgct
     2941 ggctcaccgg cgattggcag ctcgtttggg aagcgccaca gtggctcgcg gtcaacagga
     3001 ttcaggaacg gctcgcggta atggtccatc tcgacttcag tcagcgggcg gacgacaccc
     3061 atcggcagcg taccctcgat aaaaacgttc tgatcgatga tcagcttgcg gccgacgtcg
     3121 gtggtgcgga aggcctggaa ggtctcgcgg gcaaattctg gccattcgtc ccaggtcggg
     3181 atagggcgga tgaactccat aaatgcaata cctttgacgc gctctggatt gcgcttggcc
     3241 cagtggaaac ccagagcgga gccccagtcg tgaatgacca ggacgacctc ttccagaccc
     3301 agggcttcga tgaaggcatc catgaagcgg acgtggtcgt cgaagaaata acccaggtct
     3361 ggtttgtcgg atttgcccat accgatcagg tctggagcaa tgcagcgatg ggtcggtgca
     3421 acatgcggga tgatgttgcg ccacacgtag gaggaggtcg ggttaccgtg caggaacagc
     3481 acaggggtgc catcgcgcgg accaacatcg acgtagtgca tgcgctcgcc caggacttcc
     3541 acataatggg ggtcgaatgg aaagccagta ccgatttcgg atccatcgtt atcgctctga
     3601 aagtacagat cctcagtggt tggctcgaga gaaaccaagc aaagtattac tttgtctgga
     3661 cttatttcat cttcagtgtc tgggatcgtg ggcaacagag cagatcgcgt taagccccaa
     3721 aatttttgag gtgacatgtc ttttttggtg gccgtaaact tccatccaat atggcttgca
     3781 cagatcttac actgggcaac agtccaggca tacccaggaa accagctgtg ttctgtagaa
     3841 ggccggccta tcagattcaa gttgcaagcc ttatacacag taagtgtctc atgcacatat
     3901 ccatgaggat tcacataagc tgccatcggc ccacataagg ataaactgaa tatttcattt
     3961 ttggttgtta tttctgtttc ttgacattgt ttacagcaaa gggaagtaca tttattcata
     4021 atgtctaatt cacagcgaag tcgctggata gcactgccaa ttttaaggag ctgaattctc
     4081 aatacatcat caataggaag acaagcagct actctgtaag aaaaatctat tggatttgaa
     4141 ggaagagaat catcttttag attttcatcc cattcacgta gctgtttctt gattctgtcc
     4201 attaaggtct cagcatcata taaggaatac agccagcgag gccatgaagt tagatttgca
     4261 caatgaaact ttctcttctg gtatttctgc caccatttat atgaacattg gtcttctctt
     4321 gagacaggtt ttgaaggaaa tatctggcac ttattgaggg attctaattg aactgcagac
     4381 atggttgaag gcaacacaca ttcgggaaga atttgcactt tagcttgctg gattccatct
     4441 gactgtgttc ttagctcaag gactttgaac ctttgtcttc caattgcttt cactttcact
     4501 atctcaattc caaaatcctg ttcttctcga taggcatata tctctgctgt tgttccaaac
     4561 tgtgcttccc tttcctgtac attgctgtat gcaagaacag caaaggttct atctttctga
     4621 attaaattcc gcaccatact gacttcttga gggtgaaaaa gctgaagagg taatgtctgt
     4681 ccgggaatca ggatcatcat cacttgtgga agaactggaa tcacctgaca gctgtcgtca
     4741 tcgtgcaaag tcctgccatg aaattcttcc atatcagcac ctaggtatgt atgtgatgtc
     4801 ggcagactgg tgtcaaaatt tatgatgttt ggttttttgg cttctttact atcctggtct
     4861 tcaacttcca tttcatcttc ttcctcactc tctgcaggca ggagcggcag gtggttgccc
     4921 atgttgtgcg cagcgtcctg ctgatctcct tcgccggcca tggcgatcgc tagccctata
     4981 gtgagtcgta ttaatactct agccttaaga gctgtaattg aactgggagt ggacacctgt
     5041 ggagagaaag gcaaagtgga tgtcagtaag accaataggt gcctatcaga aacgcaagag
     5101 tcttctctgt ctcgacaagc ccagtttcta ttggtctcct taaacctgtc ttgtaacctt
     5161 gatacttacc tgcccagtgc ctcacgacca acttctgcag cttaagttcg agactgttgt
     5221 gtcagaagca ctgactgcgt tagcaattta actgtgataa actaccgcaa taaagcttct
     5281 agtgatctga cggttcacta aaccagctct gcttatatag acctcccacc gtacacgcct
     5341 accgcccatt tgcgtcaacg gggcggggtt attacgacat tttggaaagt cccgttgatt
     5401 ttggtgccaa aacaaactcc cattgacgtc aatggggtgg agacttggaa atccccgtga
     5461 gtcaaaccgc tatccacgcc cattggtgta ctgccaaaac cgcatcacca tggtaatagc
     5521 gatgactaat acgtagatgt actgccaagt aggaaagtcc cgtaaggtca tgtactgggc
     5581 ataatgccag gcgggccatt taccgtcatt gacgtcaata gggggcggac ttggcatatg
     5641 atacacttga tgtactgcca agtgggcagt ttaccgtaaa tactccaccc attgacgtca
     5701 atggaaagtc cctattggcg ttactatggg aacatacgtc attattgacg tcaatgggcg
     5761 ggggtcgttg ggcggtcagc caggcgggcc atttaccgta agttatgtaa cgcggaactc
     5821 catatatggg ctatgaacta atgaccccgt aattgattac tattaataac tagtcaataa
     5881 tcaatgccaa catggcggtc atattggaca tgagccaata taaatgtaca tattatgata
     5941 tagatacaac gtatgcaatg gccaatagcc aatattgatt tatgctatat aaccaatgaa
     6001 taatatggct aatggccaat attgaagatc tcgatcctct acgccggacg catcgtggcc
     6061 ggagtactgc ggtttatctc gatggctacg agggcagaca gtaagtggat ttaccataat
     6121 cccttaattg tacgcaccgc taaaacgcgt tcagcgcgat cacggcagca gacaggtaaa
     6181 aatggcaaca aaccacccga aaaactgccg caatcgcgcc tgataaattt taaccgtatg
     6241 aatacctatg caaccagagg gtacaggcca cattaccccc acttaatcca ctgaagctgc
     6301 catttttcat ggtttcacca tcccagcgaa gggccatcca gcgtgcgttc ctgtatttcc
     6361 ggactataaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     6421 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttc
//
