LOCUS       sequence-434057-        7286 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FLAG tagged human WIPI4..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7286)
  AUTHORS   Shimizu T, Tamura N, Nishimura T, Saito C, Yamamoto H, Mizushima N
  TITLE     Comprehensive analysis of autophagic functions of WIPI family 
            proteins and their implications for the pathogenesis of 
            beta-propeller associated neurodegeneration.
  JOURNAL   Hum Mol Genet. 2023 Aug 7;32(16):2623-2637. doi: 
            10.1093/hmg/ddad096.
  PUBMED    37364041
REFERENCE   2  (bases 1 to 7286)
  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"; journalName: 
            "Hum Mol Genet"; date: "2023-08-7- 7"; volume: "32"; issue: "16"; 
            pages: "2623-2637"
COMMENT     Sequence Label: pMRX-IP-hWIPI4-3×FLAG
FEATURES             Location/Qualifiers
     source          1..7286
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    49..406
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     enhancer        765..1144
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     complement(1246..1264)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1332..1436
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1437..1505,1506..2297)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1655..1674)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2461..3049
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2950..2969
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3203..3220
                     /label=L4440
                     /note="L4440 vector, forward primer"
     LTR             3350..3819
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     CDS             complement(4049..4648)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4132..4152)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     4629..4648
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     misc_feature    4666..5218
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    4668..5234
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4828..4847)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     5057..5074
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             complement(5268..5333)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     misc_feature    6470..6844
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     complement(6516..6532)
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     CDS             complement(6854..7270)
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     complement(6870..6886)
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     complement(6902..6924)
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
ORIGIN
        1 gacaaataca gaaacacagt cagacagaga caacacagaa cgatgctgca gcagacaaga
       61 cgcgcggcgc ggcttcggtc ccaaaccgaa agcaaaaatt cagacggagg cgggaactgt
      121 tttaggttct cgtctcctac cagaaccaca tatccctcct ctaagggggg tgcaccaaag
      181 agtccaaaac gatcgggatt tttggactca ggtcgggcca caaaaacggc ccccgaagtc
      241 cctgggacgt ctcccagggt tgcggccggg tgttccgaac tcgtcagttc caccacgggt
      301 ccgccagata cagagctagt tagctaacta gtaccgacgc aggcgcataa aatcagtcat
      361 agacactaga caatcggaca gacacagata agttgctggc cagcttacct cccggtggtg
      421 ggtcggtggt ccctgggcag gggtctcccg atcccggacg agcccccaaa tgaaagaccc
      481 ccgctgacgg gtagtcaatc actcagagga gaccctccca aggaacagcg agaccacaag
      541 tcggatgcaa ctgcaagagg gtttattgga tacacgggta cccgggcgac tcagtcaatc
      601 ggaggactgg cgcgccgagt gaggggttgt gggctctttt attgagctcg gggagcagaa
      661 gcgcgcgaac agaagcgaga agcgaactga ttggttagtt caaataaggc acagggtcat
      721 ttcaggtcct tggggcaccc tggaaacatc tgatggttct ctagcatggt aatagcgatg
      781 actaatacgt agatgtactg ccaagtagga aagtcccata aggtcatgta ctgggcataa
      841 tgccaggcgg gccatttacc gtcattgacg tcaatagggg gcgtacttgg catatgatac
      901 acttgatgta ctgccaagtg ggcagtttac cgtaaatact ccacccattg acgtcaatgg
      961 aaagtcccta ttggcgttac tatgggaaca tacgtcatta ttgacgtcaa tgggcggggg
     1021 tcgttgggcg gtcagccagg cgggccattt accgtaagtt atgtaacgcg gaactccata
     1081 tatgggctat gaactaatga ccccgtaatt gattactatt aataactagt caataatcaa
     1141 tgtcaacatg gcggtaatgt tggacatgag ccaatataaa tgtacatatt atgatatgga
     1201 tacaacgtat gcaatgggcc aagctcctat agtgaagacg aaagggcctc gtgatacgcc
     1261 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     1321 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     1381 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     1441 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     1501 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     1561 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     1621 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg
     1681 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     1741 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     1801 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     1861 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     1921 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     1981 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     2041 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     2101 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     2161 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     2221 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     2281 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttgc
     2341 ggccggccgc aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     2401 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     2461 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     2521 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     2581 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     2641 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     2701 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     2761 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     2821 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     2881 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     2941 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     3001 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     3061 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     3121 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     3181 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     3241 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcaactatg gccatttaat
     3301 gtaaatactt aagaaaaaaa accaaattaa ttttgataca tgctgcatgt gaaagacccc
     3361 cgctgacggg tagtcaatca ctcagaggag accctcccaa ggaacagcga gaccacaagt
     3421 cggatgcaac tgcaagaggg tttattggat acacgggtac ccgggcgact cagtcaatcg
     3481 gaggactggc gccccgagtg aggggttgtg ggctctttta ttgagctcgg ggagcagaag
     3541 cgcgcgaaca gaagcgagaa gcgaactgat tggttagttc aaataaggca cagggtcatt
     3601 tcaggtcctt ggggcaccct ggaaacatct gatggttctc tagaaactgc tgagggctgg
     3661 accgcatctg gggaccatct gttcttggcc ctgagccggg gcaggaactg cttaccacag
     3721 atatcctgtt tggcccatat tcagctgttc catctgttct tggccctgag ccggggcagg
     3781 aactgcttac cacagatatc ctgtttggcc catattcagc tgttccatct gttcctgacc
     3841 ttgatctgaa cttctctatt ctcagttatg tatttttcca tgccttgcaa aatggcgtta
     3901 cttaagctag cttgccaaac ctacaggtgg ggtctttcat tccccccttt ttctggagac
     3961 taaataaaat cttttatttt atcgtcgagc catggggtcg tgcgctcctt tcggtcgggc
     4021 gctgcgggtc gtggggcggg cggtcgactc aggcaccggg cttgcgggtc atgcaccagg
     4081 tgcgcggtcc ttcgggcacc tcgacgtcgg cggtgacggt gaagccgagc cgctcgtaga
     4141 aggggaggtt gcggggcgcg gaggtctcca ggaaggcggg caccccggcg cgctcggccg
     4201 cctccactcc ggggagcacg acggcgctgc ccagaccctt gccctggtgg tcgggcgaga
     4261 cgccgacggt ggccaggaac cacgcgggct ccttgggccg gtgcggcgcc aggaggcctt
     4321 ccatctgttg ctgcgcggcc agccgggaac cgctcaactc ggccatgcgc gggccgatct
     4381 cggcgaacac cgcccccgct tcgacgctct ccggcgtggt ccagaccgcc accgcggcgc
     4441 cgtcgtccgc gacccacacc ttgccgatgt cgagcccgac gcgcgtgagg aagagttctt
     4501 gcagctcggt gacccgctcg atgtggcggt ccgggtcgac ggtgtggcgc gtggcggggt
     4561 agtcggcgaa cgcggcggcg agggtgcgta cggcccgggg gacgtcgtcg cgggtggcga
     4621 ggcgcaccgt gggcttgtac tcggtcatgt cgacggttgt ggccatatta tcatcgtgtt
     4681 tttcaaagga aaaccacgtc cccgtggttc ggggggccta gacgtttttt taacctcgac
     4741 taaacacatg taaagcatgt gcaccgaggc cccagatcag atcccataca atggggtacc
     4801 ttctgggcat ccttcagccc cttgttgaat acgcttgagg agagccattt gactctttcc
     4861 acaactatcc aactcacaac gtggcactgg ggttgtgccg cctttgcagg tgtatcttat
     4921 acacgtggct tttggccgca gaggcacctg tcgccaggtg gggggttccg ctgcctgcaa
     4981 agggtcgcta cagacgttgt ttgtcttcaa gaaacttcca gaggaactgc ttccttcacg
     5041 acattcaaca gaccttgcat tcctttggcg agaggggaaa gacccctagg aatgctcgtc
     5101 aagaagacag ggccaggttt ccgggccctc acattgccaa aagacggcaa tatggtggaa
     5161 aataacatat agacaaacgc acaccggcct tattccaagc ggcttcggcc agtaacgtta
     5221 gggggggggg cggaatttac gtagcggccg ctctcgagaa atcattactt gtcatcgtca
     5281 tccttgtagt cgatgtcatg atctttataa tcaccgtcat ggtctttgta gtcgcctgaa
     5341 ccagagccga attcaaagtc atcatcatca cagatgtcaa ggtacacgtc gaaagcctct
     5401 ctgttgcagt ttccatcagg agtgaagaca tatttgtgga aggtcccatc tacgcagatg
     5461 gcaatgacag agttgacgtt cttggaagta ttgcgaccga aggcgcagat gcaagctgac
     5521 tcagcaggca cagtgaagct cgccaggctc cactgagagt ccacgtactg cccaatcata
     5581 ggccccacct tgcccacgcg agccagcgcg gagcggcggt tgaggcgggt atccttgaga
     5641 gcaaagatat ggacagtacc cttatcactg gaagcgcaga ggaaggagga gtcgtggctg
     5701 aagttaatgc agtagagggt ggcagggtca gtgcctcggc gcagctccac cagtttctcc
     5761 ttggattgtg tgtcaaagag gcgaataagg gtacccttct gggaggctga ggccactaca
     5821 gtgcctggct ggtttagaga cacacaggct atgtcactct gatgtgcatt gatcgtgaat
     5881 ggagcagacg aggtgccagg ctttgtgctc gccaggtcca caagttgcag actcccacac
     5941 ttgtgtcccg ggaacactag cagttgcttc tccaggctgg ggcagaggtc acagagcccc
     6001 ttggggttgt cccgggtatc aaactcaaac agctttcggg gattgtcggg gaaggagtac
     6061 acatagatgc ggttcttcag cacgatcacg atcttgtcat ggcgcatgcg cacagaaagc
     6121 actggcttgg tgaaggtgaa ctccagcacc agcttctcct tggagtcctt gccctcccgg
     6181 gcatcgtccc agatcagcac tgagatctct gagaacttgg gactactacc accgcccacc
     6241 aaggccagaa ggttggagcg gtgcagcatc tccaccaagc ccatgctgcc cacctgctcg
     6301 tggtccagat gccccttctc catcaagggc tccacgttgt agatgcgcac acctgtctcc
     6361 atggcgcagc aaaagcagct ttggtcttgg ttgaaacgca ggctggtcac tcctcgaagt
     6421 ggctgttgag tcatggatcc tttaagcttt atatcgatgg cagatccggc agtctagagg
     6481 atggtccacc cccggggtcg gcagccttca cgtgggcggc gtgtatccaa gctgcgatgc
     6541 cgtctacttt gagggcggtg ggggtggtca gcaggactgt gtaaggtcct ttccagcgag
     6601 gttctaggtt cttagtctgg tgtcggcgga cccacactgt gtcgccgact cggtaagggt
     6661 gaggtaccac cggtcggtcc agttgttctt ggtaggctgc cgccagaggt ctccagactt
     6721 cgtgctggac taagtagaga gcctgtaagt gagcttggag agaggggctg ttagtaactc
     6781 ttgtcatgtc agggtcaggg aagtttacaa ggggcggggg tgccccatat aagatctcat
     6841 atggccatat gggggcgcct agagaaggag tgagggctgg ataaagggag gatcgaggcg
     6901 gggtcgaacg aggaggttca agggggagag acggggcgga tggaggaaga ggaggcggag
     6961 gcttagggtg tacaaagggc ttgacccagg gaggggggtc aaaagccaag gcttcccagg
     7021 tcacgatgta ggggacctgg tctgggtgtc catgcgggcc aggtgaaaag accttgatct
     7081 taacctgggt gatgaggtct cggttaaagg tgccgtctcg cggccatccg acgttaaagg
     7141 ttggccattc tgcagagcag aaggtaaccc aacgtctctt cttgacatct accgactggt
     7201 tgtgagcgat ccgctcgaca tctttccagt gacctaaggt caaacttaag ggagtggtaa
     7261 cagtctggcc cgggcccata ttttca
//
