LOCUS       sequence-475111-        6692 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express AMPKa-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 6692)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6692)
  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            "Unpublished"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: AMPKa-HaloTag Fusion Vector
FEATURES             Location/Qualifiers
     source          1..6692
                     /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             2763..2783
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             2799..3683
                     /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(3771..3789)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3826..3947
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3837..3856)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3891..3910
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        4192..4549
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4192..4383
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4215..4235)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4400..4535
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        4564..4611
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4643..5437
                     /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(4697..4716)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5307..5326
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5501..5549
                     /note="synthetic polyadenylation signal"
     rep_origin      5706..6251
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     6152..6171
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6368..6385
                     /label=L4440
                     /note="L4440 vector, forward primer"
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 cgccatgcgc agactcagtt
     1081 cctggagaaa gatggcgaca gccgagaagc agaaacacga cgggcgggtg aagatcggcc
     1141 actacattct gggtgacacg ctgggggtcg gcaccttcgg caaagtgaag gttggcaaac
     1201 atgaattgac tgggcataaa gtagctgtga agatactcaa tcgacagaag attcggagcc
     1261 ttgatgtggt aggaaaaatc cgcagagaaa ttcagaacct caagcttttc aggcatcctc
     1321 atataattaa actgtaccag gtcatcagta caccatctga tattttcatg gtgatggaat
     1381 atgtctcagg aggagagcta tttgattata tctgtaagaa tggaaggctg gatgaaaaag
     1441 aaagtcggcg tctgttccaa cagatccttt ctggtgtgga ttattgtcac aggcatatgg
     1501 tggtccatag agatttgaaa cctgaaaatg tcctgcttga tgcacacatg aatgcaaaga
     1561 tagctgattt tggtctttca aacatgatgt cagatggtga atttttaaga acaagttgtg
     1621 gctcacccaa ctatgctgca ccagaagtaa tttcaggaag attgtatgca ggcccagagg
     1681 tagatatatg gagcagtggg gttattctct atgctttatt atgtggaacc cttccatttg
     1741 atgatgacca tgtgccaact ctttttaaga agatatgtga tgggatcttc tatacccctc
     1801 aatatttaaa tccttctgtg attagccttt tgaaacatat gctgcaggtg gatcccatga
     1861 agagggccac aatcaaagat atcagggaac atgaatggtt caaacaggac cttccaaaat
     1921 atctctttcc tgaggatcca tcatatagtt caaccatgat tgatgatgaa gccttaaaag
     1981 aagtatgtga aaagtttgag tgctcagaag aggaagttct cagctgtctt tacaacagaa
     2041 atcaccagga tcctttggca gttgcctacc atctcataat agataacagg agaataatga
     2101 atgaagccaa agatttctat ttggcgacaa gcccacctga ttcttttctt gatgatcatc
     2161 acctgactcg gccccatcct gaaagagtac cattcttggt tgctgaaaca ccaagggcac
     2221 gccataccct tgatgaatta aatccacaga aatccaaaca ccaaggtgta aggaaagcaa
     2281 aatggcattt aggaattaga agtcaaagtc gaccaaatga tattatggca gaagtatgta
     2341 gagcaatcaa acaattggat tatgaatgga aggttgtaaa cccatattat ttgcgtgtac
     2401 gaaggaagaa tcctgtgaca agcacttact ccaaaatgag tctacagtta taccaagtgg
     2461 atagtagaac ttatctactg gatttccgta gtattgatga tgaaattaca gaagccaaat
     2521 cagggactgc tactccacag agatcgggat cagttagcaa ctatcgatct tgccaaagga
     2581 gtgattcaga tgctgaggct caaggaaaat cctcagaagt ttctcttacc tcatctgtga
     2641 cctcacttga ctcttctcct gttgacctaa ctccaagacc tggaagtcac acaatagaat
     2701 tttttgagat gtgtgcaaat ctaattaaaa ttcttgcaca agtttctctc gagccaacca
     2761 ctgaggatct gtactttcag agcgataacg atggatccga aatcggtact ggctttccat
     2821 tcgaccccca ttatgtggaa gtcctgggcg agcgcatgca ctacgtcgat gttggtccgc
     2881 gcgatggcac ccctgtgctg ttcctgcacg gtaacccgac ctcctcctac gtgtggcgca
     2941 acatcatccc gcatgttgca ccgacccatc gctgcattgc tccagacctg atcggtatgg
     3001 gcaaatccga caaaccagac ctgggttatt tcttcgacga ccacgtccgc ttcatggatg
     3061 ccttcatcga agccctgggt ctggaagagg tcgtcctggt cattcacgac tggggctccg
     3121 ctctgggttt ccactgggcc aagcgcaatc cagagcgcgt caaaggtatt gcatttatgg
     3181 agttcatccg ccctatcccg acctgggacg aatggccaga atttgcccgc gagaccttcc
     3241 aggccttccg caccaccgac gtcggccgca agctgatcat cgatcagaac gtttttatcg
     3301 agggtacgct gccgatgggt gtcgtccgcc cgctgactga agtcgagatg gaccattacc
     3361 gcgagccgtt cctgaatcct gttgaccgcg agccactgtg gcgcttccca aacgagctgc
     3421 caatcgccgg tgagccagcg aacatcgtcg cgctggtcga agaatacatg gactggctgc
     3481 accagtcccc tgtcccgaag ctgctgttct ggggcacccc aggcgttctg atcccaccgg
     3541 ccgaagccgc tcgcctggcc aaaagcctgc ctaactgcaa ggctgtggac atcggcccgg
     3601 gtctgaatct gctgcaagaa gacaacccgg acctgatcgg cagcgagatc gcgcgctggc
     3661 tgtctactct ggagatttcc ggttaataga attctagagt cgacctgcag gcatgcaagc
     3721 tgatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca
     3781 ataactagca taaccccttg gggcggccgc ttcgagcaga catgataaga tacattgatg
     3841 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     3901 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     3961 gcattcattt tatgtttcag gttcaggggg agatgtggga ggttttttta agcaagtaaa
     4021 acctctacaa atgtggtaaa atcgaatttt aacaaaatat taacgcttac aatttcctga
     4081 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgc ggatctgcgc
     4141 agcaccatgg cctgaaataa cctctgaaag aggaacttgg ttaggtacct tctgaggcgg
     4201 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     4261 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     4321 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     4381 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     4441 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     4501 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agcttaatta
     4561 actgttgaca attaatcatc ggcatagtat atcggcatag tataatacga caaggtgagg
     4621 aactaaaccc aggaggcaga tcatgattga acaagatgga ttgcacgcag gttctccggc
     4681 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     4741 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     4801 gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac
     4861 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     4921 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     4981 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     5041 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     5101 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     5161 gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     5221 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     5281 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     5341 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     5401 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg
     5461 accgaccaag cgacgcccaa cctgccatca cgatggccgc aataaaatat ctttattttc
     5521 attacatctg tgtgttggtt ttttgtgtga atcgatagcg ataaggatcc gctcatgcga
     5581 tgataagctg tcaaacatga gaattacaac ttatatcgta tggggctgac ttcaggtgct
     5641 acatttgaag agataaattg cactgaaatc tagaaatatt ttatctgatt aataagatga
     5701 tcttcttgag atcgttttgg tctgcgcgta atctcttgct ctgaaaacga aaaaaccgcc
     5761 ttgcagggcg gtttttcgaa ggttctctga gctaccaact ctttgaaccg aggtaactgg
     5821 cttggaggag cgcagtcacc aaaacttgtc ctttcagttt agccttaacc ggcgcatgac
     5881 ttcaagacta actcctctaa atcaattacc agtggctgct gccagtggtg cttttgcatg
     5941 tctttccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cggactgaac
     6001 ggggggttcg tgcatacagt ccagcttgga gcgaactgcc tacccggaac tgagtgtcag
     6061 gcgtggaatg agacaaacgc ggccataaca gcggaatgac accggtaaac cgaaaggcag
     6121 gaacaggaga gcgcacgagg gagccgccag ggggaaacgc ctggtatctt tatagtcctg
     6181 tcgggtttcg ccaccactga tttgagcgtc agatttcgtg atgcttgtca ggggggcgga
     6241 gcctatggaa aaacggcttt gccgcggccc tctcacttcc ctgttaagta tcttcctggc
     6301 atcttccagg aaatctccgc cccgttcgta agccatttcc gctcgccgca gtcgaacgac
     6361 cgagcgtagc gagtcagtga gcgaggaagc ggaatatatc ctgtatcaca tattctgctg
     6421 acgcaccggt gcagcctttt ttctcctgcc acatgaagca cttcactgac accctcatca
     6481 gtgccaacat agtaagccag tatacactcc gctagcgctg atgtccggcg gtgcttttgc
     6541 cgttacgcac caccccgtca tgagattacg cgttttagcg gtgcgtacaa ttaagggatt
     6601 atggtaaatc cacttactgt ctgccctcgt agccatcgag ataaaccgca gtactccggc
     6661 cacgatgcgt ccggcgtaga ggatcgagat ct
//
