LOCUS       sequence-455752-        5761 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human granulin-4 with an N-terminal twin-Strep-FLAG tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5761)
  AUTHORS   Root J, Mendsaikhan A, Taylor G, Merino P, Nandy S, Wang M, Araujo 
            LT, Ryu D, Holler C, Thompson BM, Astarita G, Blain JF, Kukar T
  TITLE     Granulins rescue inflammation, lysosome dysfunction, lipofuscin, and
            neuropathology in a mouse model of progranulin deficiency.
  JOURNAL   Cell Rep. 2024 Nov 19;43(12):114985. doi: 
            10.1016/j.celrep.2024.114985.
  PUBMED    39565694
REFERENCE   2  (bases 1 to 5761)
  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: "Cell Rep.";
            date: "2024-11-19"; volume: "43(12):114985. doi"; pages: " 
            10.1016/j.celrep.2024.114985"
COMMENT     Sequence Label: pAAV hGRN4
FEATURES             Location/Qualifiers
     source          1..5761
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   4..133
                     /label=AAV2 ITR
     primer_bind     995..1017
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1079..1098
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             join(1194..1199,1200..1223,1224..1259,1260..1283)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="SAWSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     CDS             1293..1316
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    1584..2172
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1637..1657)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2055..2066)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2199..2216)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2205..2429
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        complement(2639..2968)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2653..2788
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2953..2973
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(3214..3802)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3294..3313)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3973..4764,4765..4833))
                     /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     4596..4615
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4834..4938)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5006..5024
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5062..5084)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5184..5203
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5417..5436
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        5726..5744
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     5726..5743
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 tagctgcgcg ctcgctcgct cactgaggcc gcccgggcaa agcccgggcg tcgggcgacc
       61 tttggtcgcc cggcctcagt gagcgagcga gcgcgcagag agggagtggc caactccatc
      121 actaggggtt ccttgtagtt aatgattaac ccgccatgct acttatctac gtagccatgc
      181 tctaggtacc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      241 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      301 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      361 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      421 tcatcgctat taccatggtc gaggtgagcc ccacgttctg cttcactctc cccatctccc
      481 ccccctcccc acccccaatt ttgtatttat ttatttttta attattttgt gcagcgatgg
      541 gggcgggggg gggggggggg cgcgcgcsgg gggggggggg ggggggsggg gggsggggsg
      601 gggsgrggcg gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt
      661 ttatggcgag gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag
      721 tcgctgcgcg ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc
      781 ggctctgact gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctcctccgg
      841 gctgtaatta gcgcttggtt taatgacggc ttgtttcttt tctgtggctg cgtgaaagcc
      901 ttgaggggct ccgggagggc cctttgtgcg gggggagcgg ctcggggctg tccgcggggg
      961 gacggctgcc ttcggggggg acggggcagg gcggggttcg gcttctggcg tgtgaccggc
     1021 ggctctagag cctctgctaa ccatgttcat gccttcttct ttttcctaca gctcctgggc
     1081 aacgtgctgg ttattgtgct gtctcatcat tttggcaaag aattggatca aagcttgcca
     1141 ccatgtggac tctggtgtcc tgggtcgcac tgaccgcagg actggtggct ggaagcgcat
     1201 ggtcccaccc acagttcgag aagggaggag gatctggagg aggatctgga gggagtgcat
     1261 ggtcacatcc tcagtttgag aagggagcca gtgactacaa agacgatgac gataaggacg
     1321 tgaaatgcga tatggaagtc agctgtccag atgggtatac atgctgtcga ctgcagtccg
     1381 gagcatgggg atgctgtcct ttcactcagg ccgtgtgctg tgaagaccac atccattgct
     1441 gtccagctgg ctttacctgc gatacacaga aaggcacctg tgagcagggg ccacaccagg
     1501 tcccctggat ggaaaaggca ccagctcatc tgagcctgcc agaccctcag gccctgaaaa
     1561 gatgactcga gcagcttatc gataatcaac ctctggatta caaaatttgt gaaagattga
     1621 ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt
     1681 tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt
     1741 tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg
     1801 tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg
     1861 ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc
     1921 gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaaat
     1981 catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct
     2041 tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg
     2101 ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg
     2161 ccgcctcccc gcatcgatac cgtcgactcg ctgatcagcc tcgactgtgc cttctagttg
     2221 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     2281 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     2341 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     2401 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     2461 gactagagca tggctacgta gataagtagc atggcgggtt aatcattaac tacaaggaac
     2521 ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc
     2581 gaccaaaggt cgcccgacgc ccgggcggcc tcagtgagcg agcgagcgcg cagagctttt
     2641 tgcaaaagcc taggcctcca aaaaagcctc ctcactactt ctggaatagc tcagaggccg
     2701 aggcggcctc ggcctctgca taaataaaaa aaattagtca gccatggggc ggagaatggg
     2761 cggaactggg cggagttagg ggcgggatgg gcggagttag gggcgggact atggttgctg
     2821 actaattgag atgcatgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac
     2881 acctggttgc tgactaattg agatgcatgc tttgcatact tctgcctgct ggggagcctg
     2941 gggactttcc acaccctaac tgacacacat tccacagctg cattaatgaa tcggccaacg
     3001 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     3061 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     3121 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     3181 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     3241 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     3301 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     3361 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     3421 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     3481 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     3541 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     3601 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     3661 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3721 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3781 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     3841 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     3901 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     3961 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     4021 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     4081 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     4141 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     4201 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     4261 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     4321 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     4381 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     4441 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     4501 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     4561 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     4621 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     4681 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     4741 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     4801 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     4861 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     4921 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     4981 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg
     5041 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     5101 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     5161 gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccattc
     5221 gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt
     5281 tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg
     5341 ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag
     5401 cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc
     5461 cggtgatgcc ggccacgatg cgtccggcgt agaggatctg gctagcgatg accctgctga
     5521 ttggttcgct gaccatttcc gggtgcggga cggcgttacc agaaactcag aaggttcgtc
     5581 caaccaaacc gactctgacg gcagtttacg agagagatga tagggtctgc ttcagtaagc
     5641 cagatgctac acaattaggc ttgtacatat tgtcgttaga acgcggctac aattaataca
     5701 taaccttatg tatcatacac atacgattta ggtgacacta tagaatacac ggaattaatt
     5761 c
//
