LOCUS       sequence-347150-        5636 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express Paragranulin without linker 1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5636)
  AUTHORS   Holler CJ, Taylor G, Deng Q, Kukar T
  TITLE     Intracellular Proteolysis of Progranulin Generates Stable, Lysosomal
            Granulins that Are Haploinsufficient in Patients with Frontotemporal
            Dementia Caused by GRN Mutations.
  JOURNAL   eNeuro. 2017 Aug 18;4(4). pii: eN-NWR-0100-17. doi: 
            10.1523/ENEURO.0100-17.2017. eCollection 2017 Jul-Aug.
  PUBMED    28828399
REFERENCE   2  (bases 1 to 5636)
  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: "eNeuro."; 
            date: "2017-08-18"; volume: "4(4). pii: eN-NWR-0100-17. doi"; pages:
            " 10.1523/ENEURO.0100-17.2017. eCollection 2017 Jul-Aug"
COMMENT     Sequence Label: pcDNA3.1(+)_Paragranulin-no linker
FEATURES             Location/Qualifiers
     source          1..5636
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(974..979,980..1003,1004..1039,1040..1063)
                     /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             1073..1096
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(1227..1244)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1233..1457
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1503..1931
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1590..1609)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1800..1821
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1940..1960)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1945..2274
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2125..2260
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2187..2206
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2341..3135
                     /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(2395..2414)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3005..3024
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3309..3430
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3346..3365)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3400..3419
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3479..3495)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3479..3495)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3492..3514)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3503..3519
                     /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(3527..3533,3534..3551,3552..3557))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3572..3593
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3710..3727)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3881..4469)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3961..3980)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4640..5431,5432..5500))
                     /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     5263..5282
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5501..5605)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttgcca ccatgtggac tctggtgtcc tgggtcgccc tgaccgctgg
      961 actggtggca ggatctgcct ggagtcaccc tcagttcgag aagggaggag gatctggagg
     1021 aggatcagga gggagcgcct ggtcccatcc acagtttgaa aaaggcgcta gtgactacaa
     1081 ggacgatgac gataaaacaa ggtgccccga cgggcagttc tgtcctgtcg cttgctgtct
     1141 ggatcccgga ggcgcaagct attcctgctg tcggccactg ctggattgac tcgagtctag
     1201 agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     1261 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     1321 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     1381 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     1441 tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc
     1501 ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     1561 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     1621 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     1681 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     1741 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     1801 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     1861 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     1921 taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     1981 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     2041 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     2101 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     2161 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct
     2221 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc
     2281 ccgggagctt gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc
     2341 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc
     2401 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca
     2461 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg
     2521 caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg
     2581 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag
     2641 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg
     2701 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc
     2761 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa
     2821 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac
     2881 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat
     2941 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac
     3001 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc
     3061 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt
     3121 gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc
     3181 tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg
     3241 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
     3301 cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     3361 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     3421 atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt
     3481 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     3541 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     3601 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     3661 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     3721 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     3781 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     3841 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     3901 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     3961 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     4021 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     4081 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     4141 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     4201 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     4261 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     4321 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     4381 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     4441 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     4501 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     4561 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     4621 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     4681 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     4741 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     4801 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     4861 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     4921 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     4981 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     5041 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     5101 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     5161 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     5221 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     5281 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     5341 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     5401 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     5461 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     5521 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     5581 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
