LOCUS       sequence-392580-        8889 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Human TOLL protein expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8889)
  TITLE     TREAT-AD Plasmids 2023
REFERENCE   2  (bases 1 to 8889)
  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"
COMMENT     Sequence Label: pDEST26-Nluc-hTLR4
FEATURES             Location/Qualifiers
     source          1..8889
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        15..318
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        319..522
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     473..493
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     519..543
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             632..1144
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             1175..1201
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    1211..1235
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3762..3786)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(3818..3837)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3819..3837)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3848..3865)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3848..3864)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3857..3879)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     polyA_signal    4269..4403
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4306..4325)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4360..4379
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4529..4984
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4616..4635)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4826..4847
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5101..5458
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5101..5292
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5124..5144)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5309..5444
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5509..6303
                     /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(5563..5582)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6173..6192
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6367..6415
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(6456..6475)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6575..6597
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        6721..6825
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6826..6894,6895..7686)
                     /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(7044..7063)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7857..8445
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8346..8365
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8599..8616
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        complement(8690..8794)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cattagatgc atgtcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
       61 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
      121 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      181 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      241 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      301 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
      361 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
      421 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
      481 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctcgtttagt gaaccgtcag
      541 atcgcctgga gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc
      601 agcctccgga ctctagccta ggccgcggac catggtcttc acactcgaag atttcgttgg
      661 ggactggcga cagacagccg gctacaacct ggaccaagtc cttgaacagg gaggtgtgtc
      721 cagtttgttt cagaatctcg gggtgtccgt aactccgatc caaaggattg tcctgagcgg
      781 tgaaaatggg ctgaagatcg acatccatgt catcatcccg tatgaaggtc tgagcggcga
      841 ccaaatgggc cagatcgaaa aaatttttaa ggtggtgtac cctgtggatg atcatcactt
      901 taaggtgatc ctgcactatg gcacactggt aatcgacggg gttacgccga acatgatcga
      961 ctatttcgga cggccgtatg aaggcatcgc cgtgttcgac ggcaaaaaga tcactgtaac
     1021 agggaccctg tggaacggca acaaaattat cgacgagcgc ctgatcaacc ccgacggctc
     1081 cctgctgttc cgagtaacca tcaacggagt gaccggctgg cggctgtgcg aacgcattct
     1141 ggcgggaggg ggtggatctg gagggggtgg atcttaccca tacgatgttc cagattacgc
     1201 ttctagatca acaagtttgt acaaaaaagc aggctccacc atgatgtctg cctcgcgcct
     1261 ggctgggact ctgatcccag ccatggcctt cctctcctgc gtgagaccag aaagctggga
     1321 gccctgcgtg gaggtggttc ctaatattac ttatcaatgc atggagctga atttctacaa
     1381 aatccccgac aacctcccct tctcaaccaa gaacctggac ctgagcttta atcccctgag
     1441 gcatttaggc agctatagct tcttcagttt cccagaactg caggtgctgg atttatccag
     1501 gtgtgaaatc cagacaattg aagatggggc atatcagagc ctaagccacc tctctacctt
     1561 aatattgaca ggaaacccca tccagagttt agccctggga gccttttctg gactatcaag
     1621 tttacagaag ctggtggctg tggagacaaa tctagcatct ctagagaact tccccattgg
     1681 acatctcaaa actttgaaag aacttaatgt ggctcacaat cttatccaat ctttcaaatt
     1741 acctgagtat ttttctaatc tgaccaatct agagcacttg gacctttcca gcaacaagat
     1801 tcaaagtatt tattgcacag acttgcgggt tctacatcaa atgcccctac tcaatctctc
     1861 tttagacctg tccctgaacc ctatgaactt tatccaacca ggtgcattta aagaaattag
     1921 gcttcataag ctgactttaa gaaataattt tgatagttta aatgtaatga aaacttgtat
     1981 tcaaggtctg gctggtttag aagtccatcg tttggttctg ggagaattta gaaatgaagg
     2041 aaacttggaa aagtttgaca aatctgctct agagggcctg tgcaatttga ccattgaaga
     2101 attccgatta gcatacttag actactacct cgatgatatt attgacttat ttaattgttt
     2161 gacaaatgtt tcttcatttt ccctggtgag tgtgactatt gaaagggtaa aagacttttc
     2221 ttataatttc ggatggcaac atttagaatt agttaactgt aaatttggac agtttcccac
     2281 attgaaactc aaatctctca aaaggcttac tttcacttcc aacaaaggtg ggaatgcttt
     2341 ttcagaagtt gatctaccaa gccttgagtt tctagatctc agtagaaatg gcttgagttt
     2401 caaaggttgc tgttctcaaa gtgattttgg gacaaccagc ctaaagtatt tagatctgag
     2461 cttcaatggt gttattacca tgagttcaaa cttcttgggc ttagaacaac tagaacatct
     2521 ggatttccag cattccaatt tgaaacaaat gagtgagttt tcagtattcc tatcactcag
     2581 aaacctcatt taccttgaca tttctcatac tcacaccaga gttgctttca atggcatctt
     2641 caatggcttg tccagtctcg aagtcttgaa aatggctggc aattctttcc aggaaaactt
     2701 ccttccagat atcttcacag agctgagaaa cttgaccttc ctggacctct ctcagtgtca
     2761 actggagcag ttgtctccaa cagcatttaa ctcactctcc agtcttcagg tactaaatat
     2821 gagccacaac aacttctttt cattggatac gtttccttat aagtgtctga actccctcca
     2881 ggttcttgat tacagtctca atcacataat gacttccaaa aaacaggaac tacagcattt
     2941 tccaagtagt ctagctttct taaatcttac tcagaatgac tttgcttgta cttgtgaaca
     3001 ccagagtttc ctgcaatgga tcaaggacca gaggcagctc ttggtggaag ttgaacgaat
     3061 ggaatgtgca acaccttcag ataagcaggg catgcctgtg ctgagtttga atatcacctg
     3121 tcagatgaat aagaccatca ttggtgtgtc ggtcctcagt gtgcttgtag tatctgttgt
     3181 agcagttctg gtctataagt tctattttca cctgatgctt cttgctggct gcataaagta
     3241 tggtagaggt gaaaacatct atgatgcctt tgttatctac tcaagccagg atgaggactg
     3301 ggtaaggaat gagctagtaa agaatttaga agaaggggtg cctccatttc agctctgcct
     3361 tcactacaga gactttattc ccggtgtggc cattgctgcc aacatcatcc atgaaggttt
     3421 ccataaaagc cgaaaggtga ttgttgtggt gtcccagcac ttcatccaga gccgctggtg
     3481 tatctttgaa tatgagattg ctcagacctg gcagtttctg agcagtcgtg ctggtatcat
     3541 cttcattgtc ctgcagaagg tggagaagac cctgctcagg cagcaggtgg agctgtaccg
     3601 ccttctcagc aggaacactt acctggagtg ggaggacagt gtcctggggc ggcacatctt
     3661 ctggagacga ctcagaaaag ccctgctgga tggtaaatca tggaatccag aaggaacagt
     3721 gggtacagga tgcaattggc aggaagcaac atctatcttg gacccagctt tcttgtacaa
     3781 agtggttgat cgcgtgcatg cgacgtcata gctctctccc tatagtgagt cgtattataa
     3841 gctaggcact ggccgtcgtt ttacaacgtc gtgactggga aaactgctag cttgggatct
     3901 ttgtgaagga accttacttc tgtggtgtga cataattgga caaactacct acagagattt
     3961 aaagctctaa ggtaaatata aaatttttaa gtgtataatg tgttaaacta gctgcatatg
     4021 cttgctgctt gagagttttg cttactgagt atgatttatg aaaatattat acacaggagc
     4081 tagtgattct aattgtttgt gtattttaga ttcacagtcc caaggctcat ttcaggcccc
     4141 tcagtcctca cagtctgttc atgatcataa tcagccatac cacatttgta gaggttttac
     4201 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     4261 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     4321 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     4381 atgtatctta tcatgtctgg atcgatcctg cattaatgaa tcggccaacg cgcggggaga
     4441 ggcggtttgc gtattggctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
     4501 cagttgcgca gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg
     4561 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     4621 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     4681 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     4741 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     4801 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     4861 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     4921 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca
     4981 atttcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgcg
     5041 gatctgcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt taggtacctt
     5101 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     5161 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     5221 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     5281 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     5341 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     5401 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     5461 gcttgattct tctgacacaa cagtctcgaa cttaaggcta gagccaccat gattgaacaa
     5521 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     5581 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     5641 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     5701 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     5761 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     5821 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     5881 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     5941 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     6001 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     6061 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     6121 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     6181 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     6241 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     6301 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgat
     6361 ggccgcaata aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg
     6421 atagcgataa ggatccgcgt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     6481 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     6541 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     6601 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     6661 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     6721 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     6781 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     6841 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     6901 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     6961 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     7021 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     7081 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     7141 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     7201 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     7261 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     7321 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     7381 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     7441 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     7501 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     7561 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     7621 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     7681 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     7741 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     7801 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     7861 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     7921 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     7981 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     8041 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     8101 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     8161 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     8221 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     8281 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     8341 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     8401 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     8461 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     8521 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     8581 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     8641 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gagcttgcaa ttcgcgcttt
     8701 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     8761 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     8821 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt attacgaggc
     8881 cctttcacg
//
