LOCUS       sequence-426014-        8141 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of full-length human LEO1. C-terminal HaloTag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8141)
  AUTHORS   Ackloo S, Li F, Szewczyk M, Seitova A, Loppnau P, Zeng H, Xu J, 
            Ahmad S, Arnautova YA, Baghaie AJ, Beldar S, Bolotokova A, Centrella
            PA, Chau I, Clark MA, Cuozzo JW, Dehghani-Tafti S, Disch JS, Dong A,
            Dumas A, Feng JA, Ghiabi P, Gibson E, Gilmer J, Goldman B, Green SR,
            Guie MA, Guilinger JP, Harms N, Herasymenko O, Houliston S, 
            Hutchinson A, Kearnes S, Keefe AD, Kimani SW, Kramer T, Kutera M, 
            Kwak HA, Lento C, Li Y, Liu J, Loup J, Machado RAC, Mulhern CJ, 
            Perveen S, Righetto GL, Riley P, Shrestha S, Sigel EA, Silva M, 
            Sintchak MD, Slakman BL, Taylor RD, Thompson J, Torng W, 
            Underkoffler C, von Rechenberg M, Walsh RT, Watson I, Wilson DJ, 
            Wolf E, Yadav M, Yazdi AK, Zhang J, Zhang Y, Santhakumar V, Edwards 
            AM, Barsyte-Lovejoy D, Schapira M, Brown PJ, Halabelian L, 
            Arrowsmith CH
  TITLE     A Target Class Ligandability Evaluation of WD40 Repeat-Containing 
            Proteins.
  JOURNAL   J Med Chem. 2025 Jan 23;68(2):1092-1112. doi: 
            10.1021/acs.jmedchem.4c02010. Epub 2024 Nov 4.
  PUBMED    39495097
REFERENCE   2  (bases 1 to 8141)
  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"; doi: 
            "10.1021/acs.jmedchem.4c02010"; journalName: "J Med Chem"; date: 
            "2025-01-23- 23"; volume: "68"; issue: "2"; pages: "1092-1112"
COMMENT     Sequence Label: pHTC-HaloTag_LEO1:1-666
FEATURES             Location/Qualifiers
     source          1..8141
                     /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             3095..3115
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             3131..4015
                     /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(4087..4105)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    4142..4263
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4153..4172)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4207..4226
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        4508..4865
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4508..4699
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4531..4551)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4716..4851
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4916..5710
                     /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(4970..4989)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5580..5599
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5774..5822
                     /note="synthetic polyadenylation signal"
     promoter        5978..6082
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6083..6151,6152..6943)
                     /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(6301..6320)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(7101..7689)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7181..7200)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    7805..8088
                     /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 gcgatcgctt ccgaattcac
     1081 catggcggat atggaggatc tcttcgggag cgacgccgac agcgaagctg agcgtaaaga
     1141 ttctgattct ggatctgact cagattctga tcaagagaat gctgcctctg gcagtaatgc
     1201 ctctggaagt gaaagtgatc aggatgaaag aggtgattca ggacaaccaa gtaataagga
     1261 actgtttgga gatgacagtg aggacgaggg agcttcacat catagtggta gtgataatca
     1321 ctctgaaaga tcagacaata gatcagaagc ttctgagcgt tctgaccatg aggacaatga
     1381 cccctcagat gtagatcagc acagtggatc agaagcccct aatgatgatg aagacgaagg
     1441 tcatagatcg gatggaggga gccatcattc agaagcagaa ggttctgaaa aagcacattc
     1501 agatgatgaa aaatggggca gagaagataa aagtgaccag tcagatgatg aaaagataca
     1561 aaattctgat gatgaggaga gggcacaagg atctgatgaa gataagctgc agaattctga
     1621 cgatgatgag aaaatgcaga acacagatga tgaggagagg cctcagcttt ccgatgatga
     1681 gagacaacag ctatctgagg aggaaaaggc taattctgat gatgaacggc cggtagcttc
     1741 tgataatgat gatgagaaac agaattctga tgatgaagaa caaccacagc tgtctgatga
     1801 agagaaaatg caaaattctg atgatgaaag gccacaggcc tcagatgaag aacacaggca
     1861 ttcagatgat gaagaggaac aggatcataa atcagaatct gcaagaggca gtgatagtga
     1921 agatgaagtt ttacgaatga aacgcaagaa tgcgattgca tctgattcag aagcggatag
     1981 tgacactgag gtgccaaaag ataatagtgg aaccatggat ttatttggag gtgcagatga
     2041 tatctcttca gggagtgatg gagaagacaa accacctact ccaggacagc ctgttgatga
     2101 aaatggattg cctcaggatc aacaggaaga ggagccaatt cctgagacca gaatagaagt
     2161 agaaataccc aaagtaaaca ctgatttagg aaacgactta tattttgtta aactgcccaa
     2221 ctttctcagt gtagagccca gaccttttga tcctcagtat tatgaagatg aatttgaaga
     2281 tgaagaaatg ctggatgaag aaggtagaac caggttaaaa ttaaaggtag aaaatactat
     2341 aagatggagg atacgccgag atgaagaagg aaatgaaatt aaagaaagca atgctcggat
     2401 agtcaagtgg tcagatggaa gcatgtccct gcatttaggc aatgaagtgt ttgatgtgta
     2461 caaagcccca ctgcagggcg accacaatca tctttttata agacaaggta ctggtctaca
     2521 gggacaagca gtctttaaaa cgaaactcac cttcagacct cactctacgg acagtgccac
     2581 acatagaaag atgactctgt cacttgcaga taggtgttca aagacacaga agattagaat
     2641 cttgccaatg gctggtcgtg atcctgaatg ccaacgcaca gaaatgatta agaaagaaga
     2701 agaacgtttg agggcttcca tacgtaggga atctcagcag cgccgaatga gagagaaaca
     2761 gcaccagcgg gggctgagcg ccagttacct ggaacctgat cgatacgatg aggaggagga
     2821 aggcgaggag tccatcagct tggctgccat taaaaaccga tataaagggg gcattcgaga
     2881 ggaacgagcc agaatctatt catcagacag tgatgaggga tcagaagaag ataaagctca
     2941 aagattactc aaagcaaaga aacttaccag tgatgaggaa ggtgaacctt ccggaaagag
     3001 aaaagcagaa gatgatgata aagcaaataa aaagcataag aagtatgtga tcagcgatga
     3061 agaggaagaa gatgatgatc tcgagccaac cactgaggat ctgtactttc agagcgataa
     3121 cgatggatcc gaaatcggta ctggctttcc attcgacccc cattatgtgg aagtcctggg
     3181 cgagcgcatg cactacgtcg atgttggtcc gcgcgatggc acccctgtgc tgttcctgca
     3241 cggtaacccg acctcctcct acgtgtggcg caacatcatc ccgcatgttg caccgaccca
     3301 tcgctgcatt gctccagacc tgatcggtat gggcaaatcc gacaaaccag acctgggtta
     3361 tttcttcgac gaccacgtcc gcttcatgga tgccttcatc gaagccctgg gtctggaaga
     3421 ggtcgtcctg gtcattcacg actggggctc cgctctgggt ttccactggg ccaagcgcaa
     3481 tccagagcgc gtcaaaggta ttgcatttat ggagttcatc cgccctatcc cgacctggga
     3541 cgaatggcca gaatttgccc gcgagacctt ccaggccttc cgcaccaccg acgtcggccg
     3601 caagctgatc atcgatcaga acgtttttat cgagggtacg ctgccgatgg gtgtcgtccg
     3661 cccgctgact gaagtcgaga tggaccatta ccgcgagccg ttcctgaatc ctgttgaccg
     3721 cgagccactg tggcgcttcc caaacgagct gccaatcgcc ggtgagccag cgaacatcgt
     3781 cgcgctggtc gaagaataca tggactggct gcaccagtcc cctgtcccga agctgctgtt
     3841 ctggggcacc ccaggcgttc tgatcccacc ggccgaagcc gctcgcctgg ccaaaagcct
     3901 gcctaactgc aaggctgtgg acatcggccc gggtctgaat ctgctgcaag aagacaaccc
     3961 ggacctgatc ggcagcgaga tcgcgcgctg gctgtctact ctggagattt ccggttaata
     4021 gaattggcat gcaagctgat ccggctgcta acaaagcccg aaaggaagct gagttggctg
     4081 ctgccaccgc tgagcaataa ctagcataac cccttggggc ggccgcttcg agcagacatg
     4141 ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt
     4201 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
     4261 gttaacaaca acaattgcat tcattttatg tttcaggttc agggggagat gtgggaggtt
     4321 tttttaagca agtaaaacct ctacaaatgt ggtaaaatcg aattttaaca aaatattaac
     4381 gcttacaatt tcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     4441 atacgcggat ctgcgcagca ccatggcctg aaataacctc tgaaagagga acttggttag
     4501 gtaccttctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt
     4561 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     4621 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     4681 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     4741 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     4801 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     4861 gcaaaaagct tgattcttct gacacaacag tctcgaactt aaggctagag ccaccatgat
     4921 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     4981 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     5041 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     5101 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     5161 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     5221 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     5281 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     5341 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     5401 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     5461 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     5521 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     5581 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     5641 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     5701 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     5761 tcacgatggc cgcaataaaa tatctttatt ttcattacat ctgtgtgttg gttttttgtg
     5821 tgaatcgata gcgataagga tcctctttgc gcttgcgttt tcccttgtcc agatagccca
     5881 gtagctgaca ttcatccggg gtcagcaccg tttctgcgga ctggctttct acgtaatggt
     5941 ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     6001 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     6061 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     6121 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     6181 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     6241 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt tcaaagttct
     6301 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     6361 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     6421 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     6481 caacttactt ctgacaacta tcggaggacc gaaggagcta accgcttttt tgcacaacat
     6541 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     6601 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     6661 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     6721 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     6781 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     6841 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     6901 acagatcgct gagataggtg cctcactgat taagcattgg taattcgaaa tgaccgacca
     6961 agcgacgccc aaccggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     7021 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     7081 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     7141 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     7201 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     7261 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     7321 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     7381 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     7441 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     7501 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     7561 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     7621 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     7681 ggatttcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     7741 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     7801 tagtccggaa atacaggaac gcacgctgga tggcccttcg ctgggatggt gaaaccatga
     7861 aaaatggcag cttcagtgga ttaagtgggg gtaatgtggc ctgtaccctc tggttgcata
     7921 ggtattcata cggttaaaat ttatcaggcg cgattgcggc agtttttcgg gtggtttgtt
     7981 gccattttta cctgtctgct gccgtgatcg cgctgaacgc gttttagcgg tgcgtacaat
     8041 taagggatta tggtaaatcc acttactgtc tgccctcgta gccatcgaga taaaccgcag
     8101 tactccggcc acgatgcgtc cggcgtagag gatcgagatc t
//
