LOCUS       sequence-478502-        6006 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Express CNOT7-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 6006)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6006)
  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            "Unpublished"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: CNOT7-HaloTag Fusion Vector
FEATURES             Location/Qualifiers
     source          1..6006
                     /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             1941..1961
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             1977..2861
                     /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(2949..2967)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3004..3125
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3015..3034)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3069..3088
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3370..3727
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3370..3561
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3393..3413)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3578..3713
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        3742..3789
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3821..4615
                     /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(3875..3894)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4485..4504
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4679..4727
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(4966..5554)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5046..5065)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    5670..5953
                     /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 cgccatgcca gcggcaactg
     1081 tagatcatag ccaaagaatt tgtgaagttt gggcttgcaa cttggatgaa gagatgaaga
     1141 aaattcgtca agttatccga aaatataatt acgttgctat ggacaccgag tttccaggtg
     1201 tggttgcaag acccattgga gaattcagga gcaatgctga ctatcaatac caactattgc
     1261 ggtgtaatgt agacttgtta aagataattc agctaggact gacatttatg aatgagcaag
     1321 gagaataccc tccaggaact tcaacttggc agtttaattt taaatttaat ttgacggagg
     1381 acatgtatgc ccaggactct atagagctac taacaacatc tggtatccag tttaaaaaac
     1441 atgaggagga aggaattgaa acccagtact ttgcagaact tcttatgact tctggagtgg
     1501 tcctctgtga aggggtcaaa tggttgtcat ttcatagcgg ttacgacttt ggctacttaa
     1561 tcaaaatcct aaccaactct aacttgcctg aagaagaact tgacttcttt gagatccttc
     1621 gattgttttt tcctgtcatt tatgatgtga agtacctcat gaagagctgc aaaaatctca
     1681 aaggtggatt acaggaggtg gcagaacagt tagagctgga acggatagga ccacaacatc
     1741 aggcaggatc tgattcattg ctcacaggaa tggccttttt caaaatgaga gaaatgttct
     1801 ttgaagatca tattgatgat gccaaatatt gtggtcattt gtatggcctt ggttctggtt
     1861 catcctatgt acagaatggc acagggaatg catatgaaga ggaagccaac aagcagtcag
     1921 tttctctcga gccaaccact gaggatctgt actttcagag cgataacgat ggatccgaaa
     1981 tcggtactgg ctttccattc gacccccatt atgtggaagt cctgggcgag cgcatgcact
     2041 acgtcgatgt tggtccgcgc gatggcaccc ctgtgctgtt cctgcacggt aacccgacct
     2101 cctcctacgt gtggcgcaac atcatcccgc atgttgcacc gacccatcgc tgcattgctc
     2161 cagacctgat cggtatgggc aaatccgaca aaccagacct gggttatttc ttcgacgacc
     2221 acgtccgctt catggatgcc ttcatcgaag ccctgggtct ggaagaggtc gtcctggtca
     2281 ttcacgactg gggctccgct ctgggtttcc actgggccaa gcgcaatcca gagcgcgtca
     2341 aaggtattgc atttatggag ttcatccgcc ctatcccgac ctgggacgaa tggccagaat
     2401 ttgcccgcga gaccttccag gccttccgca ccaccgacgt cggccgcaag ctgatcatcg
     2461 atcagaacgt ttttatcgag ggtacgctgc cgatgggtgt cgtccgcccg ctgactgaag
     2521 tcgagatgga ccattaccgc gagccgttcc tgaatcctgt tgaccgcgag ccactgtggc
     2581 gcttcccaaa cgagctgcca atcgccggtg agccagcgaa catcgtcgcg ctggtcgaag
     2641 aatacatgga ctggctgcac cagtcccctg tcccgaagct gctgttctgg ggcaccccag
     2701 gcgttctgat cccaccggcc gaagccgctc gcctggccaa aagcctgcct aactgcaagg
     2761 ctgtggacat cggcccgggt ctgaatctgc tgcaagaaga caacccggac ctgatcggca
     2821 gcgagatcgc gcgctggctg tctactctgg agatttccgg ttaatagaat tctagagtcg
     2881 acctgcaggc atgcaagctg atccggctgc taacaaagcc cgaaaggaag ctgagttggc
     2941 tgctgccacc gctgagcaat aactagcata accccttggg gcggccgctt cgagcagaca
     3001 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
     3061 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     3121 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag atgtgggagg
     3181 tttttttaag caagtaaaac ctctacaaat gtggtaaaat cgaattttaa caaaatatta
     3241 acgcttacaa tttcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     3301 gcatacgcgg atctgcgcag caccatggcc tgaaataacc tctgaaagag gaacttggtt
     3361 aggtaccttc tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa
     3421 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     3481 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     3541 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     3601 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     3661 cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     3721 ttgcaaaaag cttaattaac tgttgacaat taatcatcgg catagtatat cggcatagta
     3781 taatacgaca aggtgaggaa ctaaacccag gaggcagatc atgattgaac aagatggatt
     3841 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     3901 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     3961 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
     4021 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     4081 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     4141 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     4201 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     4261 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     4321 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
     4381 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     4441 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     4501 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     4561 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     4621 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg atggccgcaa
     4681 taaaatatct ttattttcat tacatctgtg tgttggtttt ttgtgtgaat cgatagcgat
     4741 aaggatcctc tttgcgcttg cgttttccct tgtccagata gcccagtagc tgacattcat
     4801 ccggggtcag caccgtttct gcggactggc tttctacccg gtatcagctc actcaaaggc
     4861 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     4921 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     4981 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     5041 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     5101 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     5161 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     5221 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     5281 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     5341 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     5401 tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     5461 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     5521 gcagcagatt acgcgcagaa aaaaaggatt tcaagaagat cctttgatct tttctacggg
     5581 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     5641 aaggatcttc acctagatcc ttttatagtc cggaaataca ggaacgcacg ctggatggcc
     5701 cttcgctggg atggtgaaac catgaaaaat ggcagcttca gtggattaag tgggggtaat
     5761 gtggcctgta ccctctggtt gcataggtat tcatacggtt aaaatttatc aggcgcgatt
     5821 gcggcagttt ttcgggtggt ttgttgccat ttttacctgt ctgctgccgt gatcgcgctg
     5881 aacgcgtttt agcggtgcgt acaattaagg gattatggta aatccactta ctgtctgccc
     5941 tcgtagccat cgagataaac cgcagtactc cggccacgat gcgtccggcg tagaggatcg
     6001 agatct
//
