LOCUS       sequence-481978-        6023 bp    DNA     circular SYN 05-AUG-2025
DEFINITION  Express HaloTag-MAP4K1 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 6023)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6023)
  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            "Unpublished"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: HaloTag-MAP4K1 Fusion Vector
FEATURES             Location/Qualifiers
     source          1..6023
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             38..832
                     /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(92..111)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     702..721
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(990..1578)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1070..1089)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    1704..1752
                     /note="synthetic polyadenylation signal"
     CDS             complement(1776..2813)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             1789..1797
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     promoter        complement(2844..3201)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2858..2993
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        3010..3201
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     3158..3178
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    3273..3354
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     3298..3317
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(3531..4043)
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     promoter        complement(5333..5351)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(5334..5351)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        5471..5774
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     5929..5949
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        join(5982..6023,1..6)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
ORIGIN
        1 tacgacaagg tgaggaacta aacccaggag gcagatcatg attgaacaag atggattgca
       61 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
      121 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
      181 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc
      241 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
      301 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc
      361 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
      421 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
      481 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
      541 cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca
      601 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
      661 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
      721 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
      781 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gattcgaaat
      841 gaccgaccaa gcgacgccca accggtatca gctcactcaa aggcggtaat acggttatcc
      901 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
      961 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     1021 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     1081 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     1141 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     1201 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     1261 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     1321 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     1381 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt
     1441 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     1501 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     1561 agaaaaaaag gatttcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     1621 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     1681 atccttttat agtccggaat cgtcacacaa aaaaccaaca cacagatgta atgaaaataa
     1741 agatatttta ttgcggccat cgtgatggct agagtctaaa ctcgacctac ctccttagcg
     1801 cgcggtcgtg tactgggccg gcggttgccg ctgtcggcca gcacctcgac gcagccgtcg
     1861 gtccatacgg ctgcgctccg gcgagcgatt tgtgtgcgac cgacggtgcc ggccccgctg
     1921 cggacgatgg catcgcagcg gccttgagcc caggcagcat cgtcgaagtt gccgtccacg
     1981 aggctctggt agagctgatc caggccgatg cgcagcatgt aggctcgcag acgagggctg
     2041 ccggccagct cgggatgccg gcgctcgaag tagcgagtct gctgctccat gcaagccagc
     2101 cagggccgcc agaagaagat gttggccacc tcgtactgac tgtccccgaa catagcttcg
     2161 gaccagtcga ttacggcggt gatgcggccg ttgtcggtca ggacgttgtt gctgccgaag
     2221 tcggcatgga cgaggtggcg cacctcggga cagtcttcgg cccacagcat cagttcgtcc
     2281 agggcttgag ctacgctggc ggacacggtg tcgtccatca cggtctgcca gtggtagaca
     2341 tggggatcag caatggcgca aatgaaatcc cgccaggttg tgtactggcc gatgccttgg
     2401 ggcccgaatg ggccgaagcc gctggtttga ctgaggtcgg cggcggcaat agcatccata
     2461 gcttcggcga caggctgtaa cacagcaggc agctctgttt cggggaggtc ttggagagta
     2521 acgccttggg cgcgtctact gatgcagtat gtcaggctct cgctgaactc gccgatgtcc
     2581 aacacttcgg ggatgggtag tgcagcgctg gcgaagtggc ggtacacgta gcggtctttg
     2641 tagaagccat cagcgcagct attcacccgc agtacatagc cgcgtccgcc gacatcgaag
     2701 ctgaaggctc ggctctcttc gccctccgac aactgcatca ggtcgctcac actgtcgaac
     2761 ttctcgatga gaaatttttc aacgctggta gcggtgagtt cgggcttctt catggtggcg
     2821 ctagtgtcag aagaatcgag ctttttgcaa aagcctaggc ctccaaaaaa gcctcctcac
     2881 tacttctgga atagctcaga ggccgaggcg gcctcggcct ctgcataaat aaaaaaaatt
     2941 agtcagccat ggggcggaga atgggcggaa ctgggcggag ttaggggcgg gatgggcgga
     3001 gttaggggcg ggactatggt tgctgactaa ttgagatgca tgctttgcat acttctgcct
     3061 gctggggagc ctggggactt tccacacctg gttgctgact aattgagatg catgctttgc
     3121 atacttctgc ctgctgggga gcctggggac tttccacacc ctaactgaca cacattccac
     3181 agctggttct ttccgcctca gaaggtacct aaccaagttc ctctttcaga ggttatttca
     3241 ggccatggtg ctgcgcaaga cgcgtgacgt cggcagtgaa aaaaatgctt tatttgtgaa
     3301 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
     3361 aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc
     3421 aagtaaaacc tctacaaatg tggtatggct gattatgatc cgcggccgca atacgtcgac
     3481 gttatcagct gacttcgtac gagagcctag gtgtttctag agtcgcggcc ttacgccaga
     3541 atgcgttcgc acagccgcca gccggtcact ccgttgatgg ttactcggaa cagcagggag
     3601 ccgtcggggt tgatcaggcg ctcgtcgata attttgttgc cgttccacag ggtccctgtt
     3661 acagtgatct ttttgccgtc gaacacggcg atgccttcat acggccgtcc gaaatagtcg
     3721 atcatgttcg gcgtaacccc gtcgattacc agtgtgccat agtgcaggat caccttaaag
     3781 tgatgatcat ccacagggta caccacctta aaaatttttt cgatctggcc catttggtcg
     3841 ccgctcagac cttcatacgg gatgatgaca tggatgtcga tcttcagccc attttcaccg
     3901 ctcaggacaa tcctttggat cggagttacg gacaccccga gattctgaaa caaactggac
     3961 acacctccct gttcaaggac ttggtccagg ttgtagccgg ctgtctgtcg ccagtcccca
     4021 acgaaatctt cgagtgtgaa gacgccgctc gagccgagag aaaccgccat ttccggcccg
     4081 ctgctctgct tcttaccgct ctgtggcggg gtgaggagcc cactggggag aggagaagcc
     4141 ctattttgtt cagacaacat ggctttcttt gctcgggctt tgtccagcaa atccaagctg
     4201 tctctgtggg tctgtatgtt gtgtgcatct tcatcagcga cgcccctgaa gtgcttcagt
     4261 tttgagctcc ccgtctccct tataaccatg gcaaatggaa ccatccactt gacacagttc
     4321 tctatgtcgc accactgata ccctgaaacc ttttgcatca attcagatga cgagaaatga
     4381 tacaaggccg aagcagcaag tataccataa ggaaattcaa ggcagtcaac atccaggaca
     4441 cagagatcca acagctctgc aatctgtata aagatttgct ggggatactg cggcagtagc
     4501 acttcatgta agtcatttag atatgcaacc tgcatgtata cattcagcca ggacacaata
     4561 gtcaggggac ttaaacgcca cttaagggcc ttcataatca ttaattccat ggtgagaatt
     4621 tcatctcctg aacaagctcc atctgtcaca tacgcaaact ggtgcaactt tggaggatag
     4681 atttcctcaa gtttggctgc aataaataaa gatgaaatcc caataagctg taaaagagtt
     4741 tttacaacat tttcttgtgt cgccatatac cggtcaaaga aatcttgtgc caagtaaaag
     4801 gtctccctgt gaagtttata gacttcacac acctccatta accaatccag aagaattgct
     4861 cgcatttttg gctgcagaag agggtgttgc tcaagaaagt gctgatccct taagtatgtc
     4921 ttttccttgt ttaacatgat tttccagact tcctctctat ttgcccagct cagtacaggc
     4981 agcggggagc ctctggatgg tgcaataatc cgaggcttgc acgttgagtt tgggtaaacc
     5041 cggtcatcat cttctttgtc aggtgtgggg atcagggagc aggggtctgc acagactgca
     5101 ttattgtccc aaggctggct cccacactgg tccctcgccg tcctgtcgat tttggccatt
     5161 tcttcatctg gatcctgcaa aaaaacggtc acgtttgcct tcctcttcct ggagcgagcc
     5221 gagaactccg cgccgccgtc ctccttcatg gtgtcccgct ccttcgcatc ccgctccctg
     5281 cgctccctcg gcatggcgat cgctagctgc aaaaagaaca agtagcttgt attctatagt
     5341 gtcacctaaa tgggcgatct gacggttcac taaaccagct ctgcttatat aggcctccct
     5401 ggcgcgccac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
     5461 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
     5521 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
     5581 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
     5641 atcatatgcc aagtccgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
     5701 atgcccagta catgacctta cgggactttc ctacttggca gtacatctac gtattagtca
     5761 tcgctattac catggtgatg cggttttggc agtacaccaa tgggcgtgga tagcggtttg
     5821 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
     5881 aaaatcaacg ggactttcca aaatgtcgta ataaccccgc cccgttgacg caaatgggcg
     5941 gtaggcgtgt acggtgggag gtcaaatcga attctaatac tgttgacaat taatcatcgg
     6001 catagtatat cggcatagta taa
//
