LOCUS       sequence-350263-        7410 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses WIPI2B (I92E) fused to Halo in mammalian cells to abrogate
            interaction with ATG16L1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7410)
  AUTHORS   Strong LM, Chang C, Riley JF, Boecker CA, Flower TG, Buffalo CZ, Ren
            X, Stavoe AK, Holzbaur EL, Hurley JH
  TITLE     Structural basis for membrane recruitment of ATG16L1 by WIPI2 in 
            autophagy.
  JOURNAL   Elife. 2021 Sep 10;10. pii: 70372. doi: 10.7554/eLife.70372.
  PUBMED    34505572
REFERENCE   2  (bases 1 to 7410)
  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: "Elife."; 
            date: "2021-09-10"; volume: "10. pii: 70372. doi"; pages: " 
            10.7554/eLife.70372"
COMMENT     Sequence Label: Halo-WIPI2B I92E
FEATURES             Location/Qualifiers
     source          1..7410
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        109..466
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        109..300
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(132..152)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      317..452
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             517..1311
                     /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(571..590)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1181..1200
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1375..1423
                     /note="synthetic polyadenylation signal"
     promoter        1579..1683
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1684..1752,1753..2544)
                     /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(1902..1921)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(2702..3290)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2782..2801)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    3406..3689
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     enhancer        3880..4259
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4260..4463
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4414..4434
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          4599..4731
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     4714..4733
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     4775..4794
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4775..4793
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      4803..4812
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4809..5699
                     /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             5712..5732
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     primer_bind     complement(7103..7121)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    7153..7274
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7164..7183)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     7218..7237
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
ORIGIN
        1 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgcgga tctgcgcagc
       61 accatggcct gaaataacct ctgaaagagg aacttggtta ggtaccttct gaggcggaaa
      121 gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
      181 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
      241 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
      301 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
      361 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt
      421 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc ttgattcttc
      481 tgacacaaca gtctcgaact taaggctaga gccaccatga ttgaacaaga tggattgcac
      541 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
      601 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
      661 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
      721 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
      781 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
      841 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
      901 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
      961 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     1021 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat
     1081 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     1141 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     1201 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     1261 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     1321 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgatgg ccgcaataaa
     1381 atatctttat tttcattaca tctgtgtgtt ggttttttgt gtgaatcgat agcgataagg
     1441 atcctctttg cgcttgcgtt ttcccttgtc cagatagccc agtagctgac attcatccgg
     1501 ggtcagcacc gtttctgcgg actggctttc tacgtaatgg tttcttagac gtcaggtggc
     1561 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     1621 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     1681 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     1741 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     1801 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     1861 cccgaagaac gttttccaat gatgagcact ttcaaagttc tgctatgtgg cgcggtatta
     1921 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     1981 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     2041 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaact
     2101 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     2161 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     2221 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     2281 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     2341 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     2401 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     2461 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     2521 gcctcactga ttaagcattg gtaattcgaa atgaccgacc aagcgacgcc caaccggtat
     2581 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     2641 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     2701 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     2761 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     2821 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     2881 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     2941 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     3001 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     3061 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     3121 ctaactacgg ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta
     3181 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     3241 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatttcaa gaagatcctt
     3301 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     3361 tcatgagatt atcaaaaagg atcttcacct agatcctttt atagtccgga aatacaggaa
     3421 cgcacgctgg atggcccttc gctgggatgg tgaaaccatg aaaaatggca gcttcagtgg
     3481 attaagtggg ggtaatgtgg cctgtaccct ctggttgcat aggtattcat acggttaaaa
     3541 tttatcaggc gcgattgcgg cagtttttcg ggtggtttgt tgccattttt acctgtctgc
     3601 tgccgtgatc gcgctgaacg cgttttagcg gtgcgtacaa ttaagggatt atggtaaatc
     3661 cacttactgt ctgccctcgt agccatcgag ataaaccgca gtactccggc cacgatgcgt
     3721 ccggcgtaga ggatcgagat cttcaatatt ggccattagc catattattc attggttata
     3781 tagcataaat caatattggc tattggccat tgcatacgtt gtatctatat cataatatgt
     3841 acatttatat tggctcatgt ccaatatgac cgccatgttg gcattgatta ttgactagtt
     3901 attaatagta atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta
     3961 cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt
     4021 caataatgac gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg
     4081 tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagtc
     4141 cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga
     4201 ccttacggga ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg
     4261 tgatgcggtt ttggcagtac accaatgggc gtggatagcg gtttgactca cggggatttc
     4321 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
     4381 ttccaaaatg tcgtaataac cccgccccgt tgacgcaaat gggcggtagg cgtgtacggt
     4441 gggaggtcta tataagcaga gctggtttag tgaaccgtca gatcactaga agctttattg
     4501 cggtagttta tcacagttaa attgctaacg cagtcagtgc ttctgacaca acagtctcga
     4561 acttaagctg cagaagttgg tcgtgaggca ctgggcaggt aagtatcaag gttacaagac
     4621 aggtttaagg agaccaatag aaactgggct tgtcgagaca gagaagactc ttgcgtttct
     4681 gataggcacc tattggtctt actgacatcc actttgcctt tctctccaca ggtgtccact
     4741 cccagttcaa ttacagctct taaggctaga gtattaatac gactcactat agggctagca
     4801 aagccaccat ggcagaaatc ggtactggct ttccattcga cccccattat gtggaagtcc
     4861 tgggcgagcg catgcactac gtcgatgttg gtccgcgcga tggcacccct gtgctgttcc
     4921 tgcacggtaa cccgacctcc tcctacgtgt ggcgcaacat catcccgcat gttgcaccga
     4981 cccatcgctg cattgctcca gacctgatcg gtatgggcaa atccgacaaa ccagacctgg
     5041 gttatttctt cgacgaccac gtccgcttca tggatgcctt catcgaagcc ctgggtctgg
     5101 aagaggtcgt cctggtcatt cacgactggg gctccgctct gggtttccac tgggccaagc
     5161 gcaatccaga gcgcgtcaaa ggtattgcat ttatggagtt catccgccct atcccgacct
     5221 gggacgaatg gccagaattt gcccgcgaga ccttccaggc cttccgcacc accgacgtcg
     5281 gccgcaagct gatcatcgat cagaacgttt ttatcgaggg tacgctgccg atgggtgtcg
     5341 tccgcccgct gactgaagtc gagatggacc attaccgcga gccgttcctg aatcctgttg
     5401 accgcgagcc actgtggcgc ttcccaaacg agctgccaat cgccggtgag ccagcgaaca
     5461 tcgtcgcgct ggtcgaagaa tacatggact ggctgcacca gtcccctgtc ccgaagctgc
     5521 tgttctgggg caccccaggc gttctgatcc caccggccga agccgctcgc ctggccaaaa
     5581 gcctgcctaa ctgcaaggct gtggacatcg gcccgggtct gaatctgctg caagaagaca
     5641 acccggacct gatcggcagc gagatcgcgc gctggctgtc gacgctcgag atttccggcg
     5701 agccaaccac tgaggatctg tactttcaga gcgataacgc gatcgcttcc gaattcggat
     5761 ccatgaacct ggcgagccag agcggggagg ccggcgccgg ccagctgctc ttcgccaact
     5821 tcaaccagga caacacgtcc ctagctgttg gtagtaagtc cggttataaa tttttctccc
     5881 tttcttctgt ggataagctg gaacagatct atgaatgcac cgatacggaa gatgtgtgca
     5941 ttgtagagag attgttctcc agcagcctag tggccatcgt gagccttaaa gcaccaagga
     6001 agctaaaggt ttgccacttt aagaagggaa ctgaggaatg caactacagc tactccaaca
     6061 cgattctggc tgtgaagctc aacaggcaga ggctgatagt atgcctggag gagtccctgt
     6121 acatccacaa cattcgggac atgaaggtgc tgcatacgat cagggagacg cctccaaacc
     6181 ctgcaggcct gtgtgcgctg tcaatcaaca acgacaactg ctacttggcg tacccaggga
     6241 gcgcgaccat cggagaggtg caggtcttcg ataccattaa tttgagagct gcaaacatga
     6301 ttccggctca cgacagtcct ttagcggcac tggcctttga cgcaagtgga actaaacttg
     6361 ccacggcttc ggagaagggg accgtgatta gggtattttc cattccagaa ggacaaaaac
     6421 tctttgagtt tcggagagga gtaaagaggt gcgtgagcat ctgctccctg gccttcagca
     6481 tggacggcat gttcctctcc gcctccagca acactgagac cgtgcacatc ttcaaactcg
     6541 agactgtgaa agaaaaaccc ccagaggagc ccaccacctg gaccgggtac ttcgggaaag
     6601 tgctcatggc ctccaccagc tacctgcctt cccaagtgac agaaatgttc aaccagggca
     6661 gagccttcgc cacggtccgc ctgccattct gcggccacaa aaacatctgc tcgctagcca
     6721 caattcagaa gatcccgcgg ttgttggtgg gtgccgccga cgggtacctg tacatgtaca
     6781 acctggaccc ccaggagggc ggcgagtgtg ccctgatgaa gcagcaccgg ctggacggca
     6841 gtctggaaac gaccaatgag atcttggact ctgcctctca cgactgcccc ttagtcactc
     6901 agacatacgg cgcagctgca ggaaaaggta cttacgtgcc ttcatcccca acgagacttg
     6961 cctacacaga cgacctgggt gctgtgggtg gcgcctgcct ggaggacgag gccagcgccc
     7021 tgcgcctgga tgaggacagc gagcacccgc ccatgattct tcggactgac tgagcggccg
     7081 cttgctgagt tggctgctgc caccgctgag caataactag cataacccct tggccgcttc
     7141 gagcagacat gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa
     7201 aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     7261 gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggaga
     7321 tgtgggaggt ttttttaagc aagtaaaacc tctacaaatg tggtaaaatc gaattttaac
     7381 aaaatattaa cgcttacaat ttcctgatgc
//
