LOCUS       sequence-434058-        7514 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FLAG tagged human WIPI2B..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7514)
  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 7514)
  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-hWIPI2B-3×FLAG
FEATURES             Location/Qualifiers
     source          1..7514
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    163..520
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     enhancer        879..1258
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     complement(1360..1378)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1446..1550
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1551..1619,1620..2411)
                     /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(1769..1788)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2575..3163
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3064..3083
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3317..3334
                     /label=L4440
                     /note="L4440 vector, forward primer"
     LTR             3464..3933
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     CDS             complement(4163..4762)
                     /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(4246..4266)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     4743..4762
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     misc_feature    4780..5332
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    4782..5348
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4942..4961)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     5171..5188
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             complement(5382..5447)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     misc_feature    6812..7186
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     complement(6858..6874)
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     CDS             complement(join(7196..7514,1..98))
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     complement(7212..7228)
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     complement(7244..7266)
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
ORIGIN
        1 cgtctcttct tgacatctac cgactggttg tgagcgatcc gctcgacatc tttccagtga
       61 cctaaggtca aacttaaggg agtggtaaca gtctggcccg ggcccatatt ttcagacaaa
      121 tacagaaaca cagtcagaca gagacaacac agaacgatgc tgcagcagac aagacgcgcg
      181 gcgcggcttc ggtcccaaac cgaaagcaaa aattcagacg gaggcgggaa ctgttttagg
      241 ttctcgtctc ctaccagaac cacatatccc tcctctaagg ggggtgcacc aaagagtcca
      301 aaacgatcgg gatttttgga ctcaggtcgg gccacaaaaa cggcccccga agtccctggg
      361 acgtctccca gggttgcggc cgggtgttcc gaactcgtca gttccaccac gggtccgcca
      421 gatacagagc tagttagcta actagtaccg acgcaggcgc ataaaatcag tcatagacac
      481 tagacaatcg gacagacaca gataagttgc tggccagctt acctcccggt ggtgggtcgg
      541 tggtccctgg gcaggggtct cccgatcccg gacgagcccc caaatgaaag acccccgctg
      601 acgggtagtc aatcactcag aggagaccct cccaaggaac agcgagacca caagtcggat
      661 gcaactgcaa gagggtttat tggatacacg ggtacccggg cgactcagtc aatcggagga
      721 ctggcgcgcc gagtgagggg ttgtgggctc ttttattgag ctcggggagc agaagcgcgc
      781 gaacagaagc gagaagcgaa ctgattggtt agttcaaata aggcacaggg tcatttcagg
      841 tccttggggc accctggaaa catctgatgg ttctctagca tggtaatagc gatgactaat
      901 acgtagatgt actgccaagt aggaaagtcc cataaggtca tgtactgggc ataatgccag
      961 gcgggccatt taccgtcatt gacgtcaata gggggcgtac ttggcatatg atacacttga
     1021 tgtactgcca agtgggcagt ttaccgtaaa tactccaccc attgacgtca atggaaagtc
     1081 cctattggcg ttactatggg aacatacgtc attattgacg tcaatgggcg ggggtcgttg
     1141 ggcggtcagc caggcgggcc atttaccgta agttatgtaa cgcggaactc catatatggg
     1201 ctatgaacta atgaccccgt aattgattac tattaataac tagtcaataa tcaatgtcaa
     1261 catggcggta atgttggaca tgagccaata taaatgtaca tattatgata tggatacaac
     1321 gtatgcaatg ggccaagctc ctatagtgaa gacgaaaggg cctcgtgata cgcctatttt
     1381 tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa
     1441 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     1501 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     1561 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     1621 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     1681 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     1741 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtgttgacg
     1801 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     1861 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     1921 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     1981 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     2041 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     2101 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     2161 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     2221 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     2281 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     2341 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     2401 agcattggta actgtcagac caagtttact catatatact ttagattgat ttgcggccgg
     2461 ccgcaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     2521 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     2581 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     2641 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     2701 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     2761 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     2821 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     2881 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     2941 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     3001 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     3061 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     3121 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     3181 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     3241 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     3301 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     3361 accgcctctc cccgcgcgtt ggccgattca ttaatgcaac tatggccatt taatgtaaat
     3421 acttaagaaa aaaaaccaaa ttaattttga tacatgctgc atgtgaaaga cccccgctga
     3481 cgggtagtca atcactcaga ggagaccctc ccaaggaaca gcgagaccac aagtcggatg
     3541 caactgcaag agggtttatt ggatacacgg gtacccgggc gactcagtca atcggaggac
     3601 tggcgccccg agtgaggggt tgtgggctct tttattgagc tcggggagca gaagcgcgcg
     3661 aacagaagcg agaagcgaac tgattggtta gttcaaataa ggcacagggt catttcaggt
     3721 ccttggggca ccctggaaac atctgatggt tctctagaaa ctgctgaggg ctggaccgca
     3781 tctggggacc atctgttctt ggccctgagc cggggcagga actgcttacc acagatatcc
     3841 tgtttggccc atattcagct gttccatctg ttcttggccc tgagccgggg caggaactgc
     3901 ttaccacaga tatcctgttt ggcccatatt cagctgttcc atctgttcct gaccttgatc
     3961 tgaacttctc tattctcagt tatgtatttt tccatgcctt gcaaaatggc gttacttaag
     4021 ctagcttgcc aaacctacag gtggggtctt tcattccccc ctttttctgg agactaaata
     4081 aaatctttta ttttatcgtc gagccatggg gtcgtgcgct cctttcggtc gggcgctgcg
     4141 ggtcgtgggg cgggcggtcg actcaggcac cgggcttgcg ggtcatgcac caggtgcgcg
     4201 gtccttcggg cacctcgacg tcggcggtga cggtgaagcc gagccgctcg tagaagggga
     4261 ggttgcgggg cgcggaggtc tccaggaagg cgggcacccc ggcgcgctcg gccgcctcca
     4321 ctccggggag cacgacggcg ctgcccagac ccttgccctg gtggtcgggc gagacgccga
     4381 cggtggccag gaaccacgcg ggctccttgg gccggtgcgg cgccaggagg ccttccatct
     4441 gttgctgcgc ggccagccgg gaaccgctca actcggccat gcgcgggccg atctcggcga
     4501 acaccgcccc cgcttcgacg ctctccggcg tggtccagac cgccaccgcg gcgccgtcgt
     4561 ccgcgaccca caccttgccg atgtcgagcc cgacgcgcgt gaggaagagt tcttgcagct
     4621 cggtgacccg ctcgatgtgg cggtccgggt cgacggtgtg gcgcgtggcg gggtagtcgg
     4681 cgaacgcggc ggcgagggtg cgtacggccc gggggacgtc gtcgcgggtg gcgaggcgca
     4741 ccgtgggctt gtactcggtc atgtcgacgg ttgtggccat attatcatcg tgtttttcaa
     4801 aggaaaacca cgtccccgtg gttcgggggg cctagacgtt tttttaacct cgactaaaca
     4861 catgtaaagc atgtgcaccg aggccccaga tcagatccca tacaatgggg taccttctgg
     4921 gcatccttca gccccttgtt gaatacgctt gaggagagcc atttgactct ttccacaact
     4981 atccaactca caacgtggca ctggggttgt gccgcctttg caggtgtatc ttatacacgt
     5041 ggcttttggc cgcagaggca cctgtcgcca ggtggggggt tccgctgcct gcaaagggtc
     5101 gctacagacg ttgtttgtct tcaagaaact tccagaggaa ctgcttcctt cacgacattc
     5161 aacagacctt gcattccttt ggcgagaggg gaaagacccc taggaatgct cgtcaagaag
     5221 acagggccag gtttccgggc cctcacattg ccaaaagacg gcaatatggt ggaaaataac
     5281 atatagacaa acgcacaccg gccttattcc aagcggcttc ggccagtaac gttagggggg
     5341 ggggcggaat ttacgtagcg gccgctctcg agaaatcatt acttgtcatc gtcatccttg
     5401 tagtcgatgt catgatcttt ataatcaccg tcatggtctt tgtagtcgcc tgaaccagag
     5461 ccgaattcgt cagtccgaag aatcatgggc gggtgctcgc tgtcctcatc caggcgcagg
     5521 gcgctggcct cgtcctccag gcaggcgcca cccacagcac ccaggtcgtc tgtgtaggca
     5581 agtctcgttg gggatgaagg cacgtaagta ccttttcctg cagctgcgcc gtatgtctga
     5641 gtgactaagg ggcagtcgtg agaggcagag tccaagatct cattggtcgt ttccagactg
     5701 ccgtccagcc ggtgctgctt catcagggca cactcgccgc cctcctgggg gtccaggttg
     5761 tacatgtaca ggtacccgtc ggcggcaccc accaacaacc gcgggatctt ctgaattgtg
     5821 gctagcgagc agatgttttt gtggccgcag aatggcaggc ggaccgtggc gaaggctctg
     5881 ccctggttga acatttctgt cacttgggaa ggcaggtagc tggtggaggc catgagcact
     5941 ttcccgaagt acccggtcca ggtggtgggc tcctctgggg gtttttcttt cacagtctcg
     6001 agtttgaaga tgtgcacggt ctcagtgttg ctggaggcgg agaggaacat gccgtccatg
     6061 ctgaaggcca gggagcagat gctcacgcac ctctttactc ctctccgaaa ctcaaagagt
     6121 ttttgtcctt ctggaatgga aaatacccta atcacggtcc ccttctccga agccgtggca
     6181 agtttagttc cacttgcgtc aaaggccagt gccgctaaag gactgtcgtg agccggaatc
     6241 atgtttgcag ctctcaaatt aatggtatcg aagacctgca cctctccgat ggtcgcgctc
     6301 cctgggtacg ccaagtagca gttgtcgttg ttgattgaca gcgcacacag gcctgcaggg
     6361 tttggaggcg tctccctgat cgtatgcagc accttcatgt cccgaatgtt gtggatgtac
     6421 agggactcct ccaggcatac tatcagcctc tgcctgttga gcttcacagc cagaatcgtg
     6481 ttggagtagc tgtagttgca gatctcagtt cccttcttaa agtggcaaac ctttagcttc
     6541 cttggtgctt taaggctcac gatggccact aggctgctgg agaacaatct ctctacaatg
     6601 cacacatctt ccgtatcggt gcattcatag atctgttcca gcttatccac agaagaaagg
     6661 gagaaaaatt tataaccgga cttactacca acagctaggg atgtgttgtc ctggttgaag
     6721 ttggcgaaga gcagctggcc ggcgccggcc tccccgctct ggctcgccag gttcatggat
     6781 cctttaagct ttatatcgat ggcagatccg gcagtctaga ggatggtcca cccccggggt
     6841 cggcagcctt cacgtgggcg gcgtgtatcc aagctgcgat gccgtctact ttgagggcgg
     6901 tgggggtggt cagcaggact gtgtaaggtc ctttccagcg aggttctagg ttcttagtct
     6961 ggtgtcggcg gacccacact gtgtcgccga ctcggtaagg gtgaggtacc accggtcggt
     7021 ccagttgttc ttggtaggct gccgccagag gtctccagac ttcgtgctgg actaagtaga
     7081 gagcctgtaa gtgagcttgg agagaggggc tgttagtaac tcttgtcatg tcagggtcag
     7141 ggaagtttac aaggggcggg ggtgccccat ataagatctc atatggccat atgggggcgc
     7201 ctagagaagg agtgagggct ggataaaggg aggatcgagg cggggtcgaa cgaggaggtt
     7261 caagggggag agacggggcg gatggaggaa gaggaggcgg aggcttaggg tgtacaaagg
     7321 gcttgaccca gggagggggg tcaaaagcca aggcttccca ggtcacgatg taggggacct
     7381 ggtctgggtg tccatgcggg ccaggtgaaa agaccttgat cttaacctgg gtgatgaggt
     7441 ctcggttaaa ggtgccgtct cgcggccatc cgacgttaaa ggttggccat tctgcagagc
     7501 agaaggtaac ccaa
//
