LOCUS       sequence-481974-        6846 bp    DNA     circular SYN 05-AUG-2025
DEFINITION  Express CNOT6-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 6846)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6846)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 5, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: CNOT6-HaloTag Fusion Vector
FEATURES             Location/Qualifiers
     source          1..6846
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1033..1052
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1033..1051
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2781..2801
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             2817..3701
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     primer_bind     complement(3789..3807)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3844..3965
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3855..3874)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3909..3928
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        4210..4567
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4210..4401
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4233..4253)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4418..4553
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        4582..4629
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4661..5455
                     /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(4715..4734)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5325..5344
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5519..5567
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(5806..6394)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5886..5905)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6510..6793
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tggtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt attaatacga ctcactatag ggctagcgat cgctttcgaa ggagatagaa
     1081 ccatgcccaa agaaaaatac gagccccctg accctcggag gatgtataca attatgtctt
     1141 ctgaggaagc agcaaatgga aagaaatccc actgggcaga gcttgaaata agtggaaaag
     1201 taagaagctt aagcgcatct ttgtggtcac taactcacct gacagctttg catttgagtg
     1261 acaattccct gtcccgaatt ccttcagaca ttgccaagct tcacaatctg gtgtatttgg
     1321 acctgtcatc taataaaatt cgtagcttac ccgcagaact cggaaacatg gtatcactca
     1381 gggagctcca tttaaataac aacctgttac gagttctacc ttttgagctg ggaaaactgt
     1441 ttcagttgca gactttaggc ctgaaaggaa atccccttac ccaggatata ttgaaccttt
     1501 atcaggaacc agatggaaca agacggctgc tgaactattt gcttgataat ttgtcaggta
     1561 ctgcaaaaag aattacaaca gaacaaccac ctccaaggtc ttggattatg ttacaagaac
     1621 cagataggac aaggccaact gccttgtttt ctgtcatgtg ctataatgtt ctttgtgata
     1681 aatatgcgac ccggcagtta tacggctact gtccatcatg ggcgctaaac tgggactaca
     1741 ggaaaaaggc cattattcaa gaaatcttga gctgcaatgc tgatatcgta agtcttcagg
     1801 aggttgaaac ggaacagtat tacagttttt ttctggtaga gctgaaagaa cgtggctata
     1861 atggattctt cagtcctaag tctagagcta ggacaatgtc agaacaagaa aggaaacatg
     1921 ttgatggctg tgcaatattc ttcaagacag aaaaatttac tttggttcag aaacacactg
     1981 ttgaatttaa tcagctagcc atggcaaatt ctgaggggtc tgaagctatg ctgaacagag
     2041 tcatgacaaa agataacatt ggggttgcag tactgctaga acttcggaag gaatcgattg
     2101 aaatgccgtc tggaaagcca catcttggaa cagaaaaaca acttattctt gtggctaacg
     2161 cccacatgca ttgggaccct gaatactctg atgtgaagtt ggtacaaact atgatgttcc
     2221 tctcagaagt gaagaacatt attgataaag cctctcgcaa cctcaaatcc agtgttttgg
     2281 gagaatttgg aactattcca cttgtgttat gtgcagatct taattctttg ccagactctg
     2341 gtgttgtaga atatttgagc acaggtggag tagaaacaaa tcacaaagac tttaaggagt
     2401 tgaggtataa tgaaagtctc acaaacttca gctgtcatgg gaagaatgga accaccaatg
     2461 gaaggatcac tcatggtttc aagttacaga gtgcctatga gagtggcctg atgccttaca
     2521 cgaattacac atttgatttc aagggtataa tagactacat tttctattct aaacctcagc
     2581 tgaacacctt aggcatcctg ggccctctgg accaccactg gctggttgag aataacatca
     2641 gtggctgccc gcaccccctc atcccctctg accacttctc actttttgca caactggagc
     2701 tcttactgcc tttcctgccc caagtcaacg gcatccacct tcctggcagg aggtacgtag
     2761 tttctctcga gccaaccact gaggatctgt actttcagag cgataacgat ggatccgaaa
     2821 tcggtactgg ctttccattc gacccccatt atgtggaagt cctgggcgag cgcatgcact
     2881 acgtcgatgt tggtccgcgc gatggcaccc ctgtgctgtt cctgcacggt aacccgacct
     2941 cctcctacgt gtggcgcaac atcatcccgc atgttgcacc gacccatcgc tgcattgctc
     3001 cagacctgat cggtatgggc aaatccgaca aaccagacct gggttatttc ttcgacgacc
     3061 acgtccgctt catggatgcc ttcatcgaag ccctgggtct ggaagaggtc gtcctggtca
     3121 ttcacgactg gggctccgct ctgggtttcc actgggccaa gcgcaatcca gagcgcgtca
     3181 aaggtattgc atttatggag ttcatccgcc ctatcccgac ctgggacgaa tggccagaat
     3241 ttgcccgcga gaccttccag gccttccgca ccaccgacgt cggccgcaag ctgatcatcg
     3301 atcagaacgt ttttatcgag ggtacgctgc cgatgggtgt cgtccgcccg ctgactgaag
     3361 tcgagatgga ccattaccgc gagccgttcc tgaatcctgt tgaccgcgag ccactgtggc
     3421 gcttcccaaa cgagctgcca atcgccggtg agccagcgaa catcgtcgcg ctggtcgaag
     3481 aatacatgga ctggctgcac cagtcccctg tcccgaagct gctgttctgg ggcaccccag
     3541 gcgttctgat cccaccggcc gaagccgctc gcctggccaa aagcctgcct aactgcaagg
     3601 ctgtggacat cggcccgggt ctgaatctgc tgcaagaaga caacccggac ctgatcggca
     3661 gcgagatcgc gcgctggctg tctactctgg agatttccgg ttaatagaat tctagagtcg
     3721 acctgcaggc atgcaagctg atccggctgc taacaaagcc cgaaaggaag ctgagttggc
     3781 tgctgccacc gctgagcaat aactagcata accccttggg gcggccgctt cgagcagaca
     3841 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
     3901 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     3961 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag atgtgggagg
     4021 tttttttaag caagtaaaac ctctacaaat gtggtaaaat cgaattttaa caaaatatta
     4081 acgcttacaa tttcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     4141 gcatacgcgg atctgcgcag caccatggcc tgaaataacc tctgaaagag gaacttggtt
     4201 aggtaccttc tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa
     4261 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     4321 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     4381 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     4441 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     4501 cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     4561 ttgcaaaaag cttaattaac tgttgacaat taatcatcgg catagtatat cggcatagta
     4621 taatacgaca aggtgaggaa ctaaacccag gaggcagatc atgattgaac aagatggatt
     4681 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     4741 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     4801 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
     4861 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     4921 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     4981 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     5041 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     5101 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     5161 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
     5221 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     5281 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     5341 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     5401 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     5461 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg atggccgcaa
     5521 taaaatatct ttattttcat tacatctgtg tgttggtttt ttgtgtgaat cgatagcgat
     5581 aaggatcctc tttgcgcttg cgttttccct tgtccagata gcccagtagc tgacattcat
     5641 ccggggtcag caccgtttct gcggactggc tttctacccg gtatcagctc actcaaaggc
     5701 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     5761 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     5821 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     5881 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     5941 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     6001 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     6061 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     6121 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     6181 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     6241 tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     6301 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     6361 gcagcagatt acgcgcagaa aaaaaggatt tcaagaagat cctttgatct tttctacggg
     6421 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     6481 aaggatcttc acctagatcc ttttatagtc cggaaataca ggaacgcacg ctggatggcc
     6541 cttcgctggg atggtgaaac catgaaaaat ggcagcttca gtggattaag tgggggtaat
     6601 gtggcctgta ccctctggtt gcataggtat tcatacggtt aaaatttatc aggcgcgatt
     6661 gcggcagttt ttcgggtggt ttgttgccat ttttacctgt ctgctgccgt gatcgcgctg
     6721 aacgcgtttt agcggtgcgt acaattaagg gattatggta aatccactta ctgtctgccc
     6781 tcgtagccat cgagataaac cgcagtactc cggccacgat gcgtccggcg tagaggatcg
     6841 agatct
//
