LOCUS       sequence-356538-        6181 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GFP-C9orf72 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6181)
  AUTHORS   Amick J, Roczniak-Ferguson A, Ferguson SM
  TITLE     C9orf72 binds SMCR8, localizes to lysosomes and regulates mTORC1 
            signaling.
  JOURNAL   Mol Biol Cell. 2016 Aug 24. pii: mbc.E16-01-0003.
  PUBMED    27559131
REFERENCE   2  (bases 1 to 6181)
  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: "Mol Biol 
            Cell. 2016 Aug 24. pii: mbc.E16-01-0003."
COMMENT     Sequence Label: pEGFP-C9orf72
FEATURES             Location/Qualifiers
     source          1..6181
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2969..3090
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3006..3025)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3060..3079
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3097..3552)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3234..3255)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3446..3465
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3579..3683
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3685..4042
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3685..3876
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3708..3728)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3893..4028
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4077..4871
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4077..4871
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4131..4150)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4741..4760
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5059..5078)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5103..5150
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5479..6067
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5968..5987
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggccggac tcagatctcg agctcaagct tcgaattctg cagtcgacgg
     1381 taccgcgggc ccatgtcgac tctttgccca ccgccatctc cagctgttgc caagacagag
     1441 attgctttaa gtggcaaatc acctttatta gcagctactt ttgcttactg ggacaatatt
     1501 cttggtccta gagtaaggca catttgggct ccaaagacag aacaggtact tctcagtgat
     1561 ggagaaataa cttttcttgc caaccacact ctaaatggag aaatccttcg aaatgcagag
     1621 agtggtgcta tagatgtaaa gttttttgtc ttgtctgaaa agggagtgat tattgtttca
     1681 ttaatctttg atggaaactg gaatggggat cgcagcacat atggactatc aattatactt
     1741 ccacagacag aacttagttt ctacctccca cttcatagag tgtgtgttga tagattaaca
     1801 catataatcc ggaaaggaag aatatggatg cataaggaaa gacaagaaaa tgtccagaag
     1861 attatcttag aaggcacaga gagaatggaa gatcagggtc agagtattat tccaatgctt
     1921 actggagaag tgattcctgt aatggaactg ctttcatcta tgaaatcaca cagtgttcct
     1981 gaagaaatag atatagctga tacagtactc aatgatgatg atattggtga cagctgtcat
     2041 gaaggctttc ttctcaatgc catcagctca cacttgcaaa cctgtggctg ttccgttgta
     2101 gtaggtagca gtgcagagaa agtaaataag atagtcagaa cattatgcct ttttctgact
     2161 ccagcagaga gaaaatgctc caggttatgt gaagcagaat catcatttaa atatgagtca
     2221 gggctctttg tacaaggcct gctaaaggat tcaactggaa gctttgtgct gcctttccgg
     2281 caagtcatgt atgctccata tcccaccaca cacatagatg tggatgtcaa tactgtgaag
     2341 cagatgccac cctgtcatga acatatttat aatcagcgta gatacatgag atccgagctg
     2401 acagccttct ggagagccac ttcagaagaa gacatggctc aggatacgat catctacact
     2461 gacgaaagct ttactcctga tttgaatatt tttcaagatg tcttacacag agacactcta
     2521 gtgaaagcct tcctggatca ggtctttcag ctgaaacctg gcttatctct cagaagtact
     2581 ttccttgcac agtttctact tgtccttcac agaaaagcct tgacactaat aaaatatata
     2641 gaagacgata cgcagaaggg aaaaaagccc tttaaatctc ttcggaacct gaagatagac
     2701 cttgatttaa cagcagaggg cgatcttaac ataataatgg ctctggctga gaaaattaaa
     2761 ccaggcctac actcttttat ctttggaaga cctttctaca ctagtgtgca agaacgagat
     2821 gttctaatga ctttttaagg gatccaccgg atctagataa ctgatcataa tcagccatac
     2881 cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa
     2941 acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa
     3001 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     3061 tggtttgtcc aaactcatca atgtatctta acgcgtaaat tgtaagcgtt aatattttgt
     3121 taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     3181 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     3241 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     3301 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     3361 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     3421 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     3481 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc
     3541 tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     3601 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     3661 aataatattg aaaaaggaag agtcctgagg cggaaagaac cagctgtgga atgtgtgtca
     3721 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3781 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3841 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     3901 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     3961 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     4021 tggaggccta ggcttttgca aagatcgatc aagagacagg atgaggatcg tttcgcatga
     4081 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     4141 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     4201 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcaag
     4261 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     4321 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     4381 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     4441 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     4501 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     4561 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg
     4621 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     4681 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     4741 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     4801 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     4861 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     4921 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     4981 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     5041 ccctaggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat
     5101 gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg
     5161 ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt ggggccaata
     5221 cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag gcccagggct
     5281 cgcagccaac gtcggggcgg caggccctgc catagcctca ggttactcat atatacttta
     5341 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     5401 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     5461 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     5521 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     5581 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     5641 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     5701 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     5761 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     5821 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     5881 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     5941 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     6001 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     6061 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     6121 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgccatgca
     6181 t
//
