LOCUS       sequence-479864-        6947 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express HaloTag-CISH 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 6947)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6947)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: HaloTag-CISH Fusion Vector
FEATURES             Location/Qualifiers
     source          1..6947
                     /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"
     regulatory      1061..1070
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1067..1957
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             1970..1990
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     primer_bind     complement(2893..2911)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2948..3069
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2959..2978)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3013..3032
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3314..3671
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3314..3505
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3337..3357)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3522..3657
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3722..4516
                     /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(3776..3795)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4386..4405
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4580..4628
                     /note="synthetic polyadenylation signal"
     promoter        4784..4888
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4889..4957,4958..5749)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5107..5126)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(5907..6495)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5987..6006)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6611..6894
                     /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 ggctagcaaa gccaccatgg cagaaatcgg
     1081 tactggcttt ccattcgacc cccattatgt ggaagtcctg ggcgagcgca tgcactacgt
     1141 cgatgttggt ccgcgcgatg gcacccctgt gctgttcctg cacggtaacc cgacctcctc
     1201 ctacgtgtgg cgcaacatca tcccgcatgt tgcaccgacc catcgctgca ttgctccaga
     1261 cctgatcggt atgggcaaat ccgacaaacc agacctgggt tatttcttcg acgaccacgt
     1321 ccgcttcatg gatgccttca tcgaagccct gggtctggaa gaggtcgtcc tggtcattca
     1381 cgactggggc tccgctctgg gtttccactg ggccaagcgc aatccagagc gcgtcaaagg
     1441 tattgcattt atggagttca tccgccctat cccgacctgg gacgaatggc cagaatttgc
     1501 ccgcgagacc ttccaggcct tccgcaccac cgacgtcggc cgcaagctga tcatcgatca
     1561 gaacgttttt atcgagggta cgctgccgat gggtgtcgtc cgcccgctga ctgaagtcga
     1621 gatggaccat taccgcgagc cgttcctgaa tcctgttgac cgcgagccac tgtggcgctt
     1681 cccaaacgag ctgccaatcg ccggtgagcc agcgaacatc gtcgcgctgg tcgaagaata
     1741 catggactgg ctgcaccagt cccctgtccc gaagctgctg ttctggggca ccccaggcgt
     1801 tctgatccca ccggccgaag ccgctcgcct ggccaaaagc ctgcctaact gcaaggctgt
     1861 ggacatcggc ccgggtctga atctgctgca agaagacaac ccggacctga tcggcagcga
     1921 gatcgcgcgc tggctgtcga cgctcgagat ttccggcgag ccaaccactg aggatctgta
     1981 ctttcagagc gataacgcga tcgccatggt tctctgcgtt cagggacctc gtcctttgct
     2041 ggctgttgag cggactggtc agcggccact gtgggctccg tccctggaac tgcccaagcc
     2101 agtcatgcag ccattgcctg ctggtgcctt cctcgaagag gttgcagagg gtaccccagc
     2161 ccagacagag agtgagccaa aggtgctgga cccagaggag gatctgctgt gcatagccaa
     2221 gaccttctcc taccttcggg aatctggctg gtattggggt tccattacgg ccagcgaggc
     2281 ccgacaacac ctgcagaaga tgccagaagg cacgttctta gtacgtgaca gcacgcaccc
     2341 cagctacctg ttcacgctgt cagtgaaaac cactcgtggc ccaaccaatg tacgcattga
     2401 gtatgccgac tccagcttcc gtctggactc caactgcttg tccaggccac gcatcctggc
     2461 ctttccggat gtggtcagcc ttgtgcagca ctatgtggcc tcctgcactg ctgatacccg
     2521 aagcgacagc ccagatcctg ctcccacccc agccctgcct atgcctaagg aggatgcgcc
     2581 tagtgaccca gcactgcctg ctcctccacc agctactgct gtacacctaa aactggtgca
     2641 gccctttgta cgcagaagca gtgcacgcag cctgcaacac ctgtgccgcc ttgttatcaa
     2701 ccgtctggtg gccgatgttg actgcctgcc actgccccgg cgcatggctg actacctccg
     2761 acagtacccc ttccagctcg tttaaacgaa ttcgggctcg gtacccgggg atcctctaga
     2821 gtcgacctgc aggcatgcaa gctgatccgg ctgctaacaa agcccgaaag gaagctgagt
     2881 tggctgctgc caccgctgag caataactag cataacccct tggggcggcc gcttcgagca
     2941 gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa
     3001 tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat
     3061 aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggagatgtgg
     3121 gaggtttttt taagcaagta aaacctctac aaatgtggta aaatcgaatt ttaacaaaat
     3181 attaacgctt acaatttcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     3241 caccgcatac gcggatctgc gcagcaccat ggcctgaaat aacctctgaa agaggaactt
     3301 ggttaggtac cttctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg
     3361 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     3421 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3481 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     3541 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     3601 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     3661 gcttttgcaa aaagcttgat tcttctgaca caacagtctc gaacttaagg ctagagccac
     3721 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt
     3781 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc
     3841 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact
     3901 gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt
     3961 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca
     4021 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
     4081 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
     4141 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
     4201 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga
     4261 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
     4321 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
     4381 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
     4441 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
     4501 tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac
     4561 ctgccatcac gatggccgca ataaaatatc tttattttca ttacatctgt gtgttggttt
     4621 tttgtgtgaa tcgatagcga taaggatcct ctttgcgctt gcgttttccc ttgtccagat
     4681 agcccagtag ctgacattca tccggggtca gcaccgtttc tgcggactgg ctttctacgt
     4741 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     4801 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     4861 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     4921 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     4981 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     5041 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcactttcaa
     5101 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     5161 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     5221 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     5281 tgcggccaac ttacttctga caactatcgg aggaccgaag gagctaaccg cttttttgca
     5341 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     5401 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     5461 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     5521 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     5581 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     5641 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     5701 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaat tcgaaatgac
     5761 cgaccaagcg acgcccaacc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5821 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5881 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5941 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     6001 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     6061 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     6121 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     6181 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     6241 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     6301 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt
     6361 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     6421 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     6481 aaaaaaggat ttcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     6541 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     6601 cttttatagt ccggaaatac aggaacgcac gctggatggc ccttcgctgg gatggtgaaa
     6661 ccatgaaaaa tggcagcttc agtggattaa gtgggggtaa tgtggcctgt accctctggt
     6721 tgcataggta ttcatacggt taaaatttat caggcgcgat tgcggcagtt tttcgggtgg
     6781 tttgttgcca tttttacctg tctgctgccg tgatcgcgct gaacgcgttt tagcggtgcg
     6841 tacaattaag ggattatggt aaatccactt actgtctgcc ctcgtagcca tcgagataaa
     6901 ccgcagtact ccggccacga tgcgtccggc gtagaggatc gagatct
//
