LOCUS       sequence-260947-        6063 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human NFL containing the Charcot-Marie-Tooth disease type 
            2E N98S mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6063)
  AUTHORS   Stone EJ, Uchida A, Brown A
  TITLE     Charcot-Marie-Tooth disease Type 2E/1F mutant neurofilament proteins
            assemble into neurofilaments.
  JOURNAL   Cytoskeleton (Hoboken). 2019 Jul;76(7-8):423-439. doi: 
            10.1002/cm.21566. Epub 2019 Nov 6.
  PUBMED    31574566
REFERENCE   2  (bases 1 to 6063)
  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: 
            "Cytoskeleton (Hoboken)."; date: "2019-07"; volume: 
            "76(7-8):423-439. doi"; pages: " 10.1002/cm.21566. Epub 2019 Nov 6"
COMMENT     Sequence Label: phNFL-N98S
FEATURES             Location/Qualifiers
     source          1..6063
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        159..462
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        463..666
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     617..637
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    707..763
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    2949..3070
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2986..3005)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3040..3059
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3077..3532)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3214..3235)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3426..3445
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3559..3663
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3665..4022
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3665..3856
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3688..3708)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3873..4008
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4057..4851
                     /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             4057..4851
                     /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(4111..4130)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4721..4740
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5039..5058)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5083..5130
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5459..6047
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5948..5967
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
       61 cccctgattc tgtggataac cgtattaccg ccatgcatta gttattaata gtaatcaatt
      121 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
      181 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
      241 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
      301 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
      361 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
      421 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
      481 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
      541 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
      601 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
      661 agagctggtt tagtgaaccg tcagatccgc tagcgctacc ggactcagat ctcgagctca
      721 agcttcgaat tctgcagtcg acggtaccgc gggcccggga tccgggtggc agaacggcgc
      781 cgagccgcac acagccatcc atcctccccc ttccctctct cccctgtcct ctctctccgg
      841 gctcccaccg ccgccgcggg ccggggagcc accggccgcc accatgagtt ccttcagcta
      901 cgagccgtac tactcgacct cctacaagcg gcgctacgtg gagacgcccc gggtgcacat
      961 ctccagcgtg cgcagcggct acagcaccgc acgctcagct tactccagct actcggcgcc
     1021 ggtgtcttcc tcgctgtccg tgcgccgcag ctactcctcc agctctggat cgttgatgcc
     1081 cagtctggag aacctcgacc tgagccaggt agccgccatc agcaacgacc tcaagtccat
     1141 ccgcacgcag gagaaggcgc agctccagga cctcagtgac cgcttcgcca gcttcatcga
     1201 gcgcgtgcac gagctggagc agcagaacaa ggtcctggaa gccgagctgc tggtgctgcg
     1261 ccagaagcac tccgagccat cccgcttccg ggcgctgtac gagcaggaga tccgcgacct
     1321 gcgcctggcg gcggaagatg ccaccaacga gaagcaggcg ctccagggcg agcgcgaagg
     1381 gctggaggag accctgcgca acctgcaggc gcgctatgaa gaggaggtgc tgagccgcga
     1441 ggacgccgag ggccggctga tggaagcgcg caaaggcgcc gacgaggcgg cgctcgctcg
     1501 cgccgagctc gagaagcgca tcgacagctt gatggacgaa atctcttttc tgaagaaagt
     1561 gcacgaagag gagatcgccg aactgcaggc gcagatccag tacgcgcaga tctccgtgga
     1621 gatggacgtg accaagcccg acctttccgc cgcgctcaag gacatccgcg cgcagtacga
     1681 gaagctggcc gccaagaaca tgcagaacgc tgaggaatgg ttcaagagcc gcttcaccgt
     1741 gctgaccgag agcgccgcca agaacaccga cgccgtgcgc gccgccaagg acgaggtgtc
     1801 cgagagccgt cgtctgctca aggccaagac cctggaaatc gaagcatgcc ggggcatgaa
     1861 tgaagcgctg gagaagcagc tgcaggagct ggaggacaag cagaacgccg acatcagcgc
     1921 tatgcaggac acgatcaaca aattagaaaa tgaattgagg accacaaaga gtgaaatggc
     1981 acgataccta aaagaatacc aagacctcct caacgtgaag atggctttgg atattgagat
     2041 tgcagcttac aggaaactct tggaaggcga ggagacccga ctcagtttca ccagcgtggg
     2101 aagcataacc agtggctact cccagagctc ccaggtcttt ggccgatctg cctacggcgg
     2161 tttacagacc agctcctatc tgatgtccac ccgctccttc ccgtcctact acaccagcca
     2221 tgtccaagag gagcagatcg aagtggagga aaccattgag gctgccaagg ctgaggaagc
     2281 caaggatgag cccccctctg aaggagaagc cgaggaggag gagaaggaca aggaagaggc
     2341 cgaggaagag gaggcagctg aagaggaaga agctgccaag gaagagtctg aagaagcaaa
     2401 agaagaagaa gaaggaggtg aaggtgaaga aggagaggaa accaaagaag ctgaagagga
     2461 ggagaagaaa gttgaaggtg ctggggagga acaagcagct aagaagaaag attgaacccc
     2521 catttcctta attatttcag gaataattct cccgaaatca ggtcaacccc atcaccaacc
     2581 aaccaaccag ttgagttcca gattctatgt gaattaaaaa gtcaatatat gtataattct
     2641 gagatgactt aggttggaca ttcaatgttg tgctatgaat ttcctcttta tgcagagtat
     2701 ctgtttgctt gcagagtggc tttctggctt gctgccagcc tgtgcatggt ccacaagccg
     2761 aattcagatc tggtaccacg cgtatcgata agcttgcatg cctgcaggtc gactctagag
     2821 gatccaccgg atctagataa ctgatcataa tcagccatac cacatttgta gaggttttac
     2881 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     2941 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     3001 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     3061 atgtatctta acgcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt
     3121 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca
     3181 aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta
     3241 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
     3301 cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
     3361 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
     3421 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg
     3481 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc
     3541 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     3601 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     3661 agtcctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     3721 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     3781 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     3841 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
     3901 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
     3961 cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
     4021 aagatcgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     4081 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     4141 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     4201 gtcaagaccg acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg
     4261 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     4321 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     4381 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     4441 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     4501 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     4561 gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat
     4621 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     4681 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     4741 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     4801 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     4861 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     4921 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     4981 tcctccagcg cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg
     5041 aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga
     5101 ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc
     5161 actctgtcga taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt
     5221 ccccacccca ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg
     5281 caggccctgc catagcctca ggttactcat atatacttta gattgattta aaacttcatt
     5341 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     5401 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     5461 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     5521 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     5581 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     5641 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     5701 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     5761 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     5821 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     5881 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     5941 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     6001 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     6061 cgg
//
