LOCUS       sequence-376913-        6256 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of wipi2d mutant.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6256)
  AUTHORS   Fracchiolla D, Chang C, Hurley JH, Martens S
  TITLE     A PI3K-WIPI2 positive feedback loop allosterically activates LC3 
            lipidation in autophagy.
  JOURNAL   J Cell Biol. 2020 Jul 6;219(7). pii: 151802. doi: 
            10.1083/jcb.201912098.
  PUBMED    32437499
REFERENCE   2  (bases 1 to 6256)
  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: "J Cell 
            Biol."; date: "2020-07-6"; volume: "219(7). pii: 151802. doi"; 
            pages: " 10.1083/jcb.201912098"
COMMENT     Sequence Label: pCAG-WIPI2d (FRRG/AAAA)-TS
FEATURES             Location/Qualifiers
     source          1..6256
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..663
                     /label=chicken beta-actin promoter
     intron          664..1680
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1604..1626
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1688..1707
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             3041..3061
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     regulatory      3066..3075
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3077..3100
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             3137..3160
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(3278..3297)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3343..3398
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3397..3416)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3759..3775)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3759..3775)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3772..3794)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3783..3799
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3807..3813,3814..3831,3832..3837))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3852..3873
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3932..4127
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3978..4113
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4133..4267
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4170..4189)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4224..4243
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4335..4352)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4506..5094)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4586..4605)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5265..6056,6057..6125))
                     /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     5888..5907
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6126..6230)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggggcgc gcgccaggcg gggcggggcg gggcgagggg
      541 cggggcgggg cgaggcggag aggtgcggcg gcagccaatc agagcggcgc gctccgaaag
      601 tttcctttta tggcgaggcg gcggcggcgg cggccctata aaaagcgaag cgcgcggcgg
      661 gcgggagtcg ctgcgcgctg ccttcgcccc gtgccccgct ccgccgccgc ctcgcgccgc
      721 ccgccccggc tctgactgac cgcgttactc ccacaggtga gcgggcggga cggcccttct
      781 cctccgggct gtaattagcg cttggtttaa tgacggcttg tttcttttct gtggctgcgt
      841 gaaagccttg aggggctccg ggagggccct ttgtgcgggg ggagcggctc ggggggtgcg
      901 tgcgtgtgtg tgtgcgtggg gagcgccgcg tgcggctccg cgctgcccgg cggctgtgag
      961 cgctgcgggc gcggcgcggg gctttgtgcg ctccgcagtg tgcgcgaggg gagcgcggcc
     1021 gggggcggtg ccccgcggtg cggggggggc tgcgagggga acaaaggctg cgtgcggggt
     1081 gtgtgcgtgg gggggtgagc agggggtgtg ggcgcgtcgg tcgggctgca accccccctg
     1141 cacccccctc cccgagttgc tgagcacggc ccggcttcgg gtgcggggct ccgtacgggg
     1201 cgtggcgcgg ggctcgccgt gccgggcggg gggtggcggc aggtgggggt gccgggcggg
     1261 gcggggccgc ctcgggccgg ggagggctcg ggggaggggc gcggcggccc ccggagcgcc
     1321 ggcggctgtc gaggcgcggc gagccgcagc cattgccttt tatggtaatc gtgcgagagg
     1381 gcgcagggac ttcctttgtc ccaaatctgt gcggagccga aatctgggag gcgccgccgc
     1441 accccctcta gcgggcgcgg ggcgaagcgg tgcggcgccg gcaggaagga aatgggcggg
     1501 gagggccttc gtgcgtcgcc gcgccgccgt ccccttctcc ctctccagcc tcggggctgt
     1561 ccgcgggggg acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt
     1621 gtgaccggcg gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag
     1681 ctcctgggca acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attcctcgat
     1741 cgacggtatc gatgccacca tgaacctggc gagccagagc ggggaggccg gcgccggcca
     1801 gctgctcttc gccaacttca accaggacaa cacgtcccta gctgttggta gtaagtccgg
     1861 ttataaattt ttctcccttt cttctgtgga taagctggaa cagatctatg aatgcaccga
     1921 tacggaagat gtgtgcattg tagagagatt gttctccagc agcctagtgg ccatcgtgag
     1981 ccttaaagca ccaaggaagc taaaggtttg ccactttaag aagggaactg agatctgcaa
     2041 ctacagctac tccaacacga ttctggctgt gaagctcaac aggcagaggc tgatagtatg
     2101 cctggaggag tccctgtaca tccacaacat tcgggacatg aaggtgctgc atacgatcag
     2161 ggagacgcct ccaaaccctg caggcctgtg tgcgctgtca atcaacaacg acaactgcta
     2221 cttggcgtac ccagggagcg cgaccatcgg agaggtgcag gtcttcgata ccattaattt
     2281 gagagctgca aacatgattc cggctcacga cagtccttta gcggcactgg cctttgacgc
     2341 aagtggaact aaacttgcca cggcttcgga gaaggggacc gtgattaggg tattttccat
     2401 tccagaagga caaaaactct ttgaggctgc ggcagcagta aagaggtgcg tgagcatctg
     2461 ctccctggcc ttcagcatgg acggcatgtt cctctccgcc tccagcaaca ctgagaccgt
     2521 gcacatcttc aaactcgaga ctgtgaaaga aaaaccccca gaggagccca ccacctggac
     2581 cgggtacttc gggaaagtgc tcatggcctc caccagctac ctgccttccc aagtgacaga
     2641 aatgttcaac cagggcagag ccttcgccac ggtccgcctg ccattctgcg gccacaaaaa
     2701 catctgctcg ctagccacaa ttcagaagat cccgcggttg ttggtgggtg ccgccgacgg
     2761 gtacctgtac atgtacaacc tggaccccca ggagggcggc gagtgtgccc tgatgaagca
     2821 gcaccggctg gacggcagtc tggaaacgac caatgagatc ttggactctg cctctcacga
     2881 ctgcccctta gtcactcaga catacggcgc agctgcagga aaagcctaca cagacgacct
     2941 gggtgctgtg ggtggcgcct gcctggagga cgaggccagc gccctgcgcc tggatgagga
     3001 cagcgagcac ccgcccatga ttcttcggac tgacggatcg gaaaacctgt attttcaggg
     3061 tggtaccatg gcggcctggt cacacccgca gttcgagaaa ggtggaggtg cccgaggtgg
     3121 atcgggaggt ggatcgtgga gccacccgca gttcgaaaaa tgacccgggt gatatcttaa
     3181 ttaattaact cgaggaattc actcctcagg tgcaggctgc ctatcagaag gtggtggctg
     3241 gtgtggccaa tgccctggct cacaaatacc actgagatct ttttccctct gccaaaaatt
     3301 atggggacat catgaagccc cttgagcatc tgacttctgg ctaataaagg aaatttattt
     3361 tcattgcaat agtgtgttgg aattttttgt gtctctcact cggaaggaca tatgggaggg
     3421 caaatcattt aaaacatcag aatgagtatt tggtttagag tttggcaaca tatgcccata
     3481 tgctggctgc catgaacaaa ggttggctat aaagaggtca tcagtatatg aaacagcccc
     3541 ctgctgtcca ttccttattc catagaaaag ccttgacttg aggttagatt ttttttatat
     3601 tttgttttgt gttatttttt tctttaacat ccctaaaatt ttccttacat gttttactag
     3661 ccagattttt cctcctctcc tgactactcc cagtcatagc tgtccctctt ctcttatgga
     3721 gatccctcga cctgcagccc aagcttggcg taatcatggt catagctgtt tcctgtgtga
     3781 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     3841 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     3901 cagtcgggaa acctgtcgtg ccagcggatc cgcatctcaa ttagtcagca accatagtcc
     3961 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     4021 atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat
     4081 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctaacttgtt
     4141 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4201 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     4261 ctggatccgc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     4321 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     4381 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     4441 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     4501 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     4561 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     4621 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     4681 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     4741 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     4801 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     4861 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     4921 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     4981 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     5041 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     5101 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     5161 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     5221 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     5281 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     5341 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     5401 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     5461 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     5521 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     5581 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     5641 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     5701 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     5761 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     5821 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     5881 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     5941 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     6001 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     6061 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     6121 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     6181 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     6241 cgaaaagtgc cacctg
//
