LOCUS       sequence-38889-e        7464 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression construct driving the expression of a 
            fluorescent fusion protein consisting of EGFP linked to the 
            N-terminus of rat neurofilament protein M.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7464)
  AUTHORS   Wang L, Ho CL, Sun D, Liem RK, Brown A
  TITLE     Rapid movement of axonal neurofilaments interrupted by prolonged 
            pauses.
  JOURNAL   Nat Cell Biol. 2000 Mar;2(3):137-41.
  PUBMED    10707083
REFERENCE   2  (bases 1 to 7464)
  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: "Nat Cell 
            Biol."; date: "2000-03"; volume: "2(3)"; pages: "137-41"
COMMENT     Sequence Label: pEGFP-rNFM
FEATURES             Location/Qualifiers
     source          1..7464
                     /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(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     RBS             2854..2865
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     polyA_signal    4252..4373
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4289..4308)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4343..4362
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4380..4835)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4517..4538)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4729..4748
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4862..4966
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4968..5325
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4968..5159
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4991..5011)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5176..5311
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5360..6154
                     /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             5360..6154
                     /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(5414..5433)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6024..6043
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6342..6361)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6386..6433
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6762..7350
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7251..7270
                     /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 ccggactcag atctcgagct caagcttcga attctgcagt cgacggtacc
     1381 gcgggcccaa ttccggcctc caagatgagc tacacgctgg actcgctggg caacccgtcc
     1441 gcctaccggc gggtcaccga gacccgctcc agcttcagtc gtgtgagcgg ttccccgtcc
     1501 agcggcttcc gctcgcagtc ctggtcccgc ggctcgccca gcaccgtgtc ctcctcctac
     1561 aagcgcagcg cgctcgcccc gcgcctcgcc tacagctcgg ctatgctcag ctcggccgag
     1621 agcagcctcg acttcagcca gtcctcttcg ctgcttaacg gcggctccgg cggcgactac
     1681 aagctgtccc gctccaacga gaaagagcag ctgcaggggc tgaacgaccg cttcgccggc
     1741 tacatcgaga aagtgcacta cttggaacaa cagaacaagg agatcgaggc agagatccac
     1801 gcgctgcggc agaagcaggc ctcgcacgcc cagctgggtg acgcttacga ccaggagatc
     1861 cgagagctgc gcgccaccct ggagatggtg aatcacgaga aggctcaagt gcagctggac
     1921 tctgatcact tggaggaaga catccaccgg ctcaaggagc gcttcgagga ggaggcgcgg
     1981 ctgcgggacg acaccgaggc tgccatccgg gcgctgcgca aagacataga ggagtcgtcg
     2041 atggttaagg tggagctgga caagaaggtg cagtcgctgc aggatgaggt ggccttcctg
     2101 cggagcaatc acgaagagga ggtggccgac ctgctggccc agatccaggc gtcgcacatc
     2161 accgtagagc gcaaagacta cctgaagaca gacatctcca cggcgctgaa agagatccgc
     2221 tcccagctcg agtgtcactc cgaccagaac atgcaccagg ccgaagagtg gttcaaatgc
     2281 cgctacgcca agctcaccga ggcggccgag cagaacaagg aggccatccg ctccgctaaa
     2341 gaagagatcg ccgagtaccg gcgccagctg cagtccaaga gcattgagct cgagtcggtg
     2401 cgaggcacta aggagtccct ggaacggcag ctcagcgaca tcgaggagcg ccacaaccac
     2461 gacctcagca gctaccagga caccatccag cagctggaaa atgagcttcg gggaacaaag
     2521 tgggaaatgg ctcgtcattt gcgagaatac caggatctcc ttaacgtcaa gatggctctg
     2581 gacatcgaga tcgccgcata taggaaacta ctggagggtg aagagaccag atttagcaca
     2641 ttttcaggaa gcatcactgg gcctctgtac acacaccgac agccctcagt cacaatatcc
     2701 agtaagattc agaagaccaa agtcgaggcc cccaagctca aggtccaaca caaatttgtg
     2761 gaggagatca ttgaggagac taaagtggaa gatgagaagt cagaaatgga agacgccctc
     2821 acagtcattg cagaggaatt ggcagcctct gccaaagagg agaaagaaga ggcagaagaa
     2881 aaggaagagg aaccggaagt tgaaaagtct cccgtgaagt ctcctgaggc taaggaagag
     2941 gaggaagggg aaaaggagga agaagaggaa ggccaagagg aagaagagga ggaagatgaa
     3001 ggtgtcaagt cagaccaggc agaagaggga ggatctgaga aggaaggctc gagtgaaaag
     3061 gatgaaggtg agcaagaaga agaaggggaa actgaggcag aaggtgaagg agaggaagca
     3121 gaagctaagg aggaaaagaa aacagaggga aaggtcgagg aaatggctat caaggaggaa
     3181 atcaaggtcg agaagcccga gaaagccaag tcccctgtgc caaaatcacc cgtggaagaa
     3241 gtaaagccaa aaccagaagc caaagccgga aaggatgagc agaaggagga agagaaagtt
     3301 gaggagaaga aggaggtagc caaggaatca cccaaggaag agaaggtgga gaaaaaggag
     3361 gagaagccaa aagatgtccc agataaaaag aaggctgagt ccccagtgaa agaaaaggcc
     3421 gtagaggaaa tgatcaccat tactaagtcg gtaaaggtga gcctggagaa agacaccaaa
     3481 gaggagaagc ctcagcagca ggagaaggtg aaggagaagg cagaggagga ggggggtagt
     3541 gaggaggaag tgggtgacaa aagcccgcaa gaatccaaga aggaagacat agctatcaat
     3601 ggggaggtgg aaggaaaaga ggaggaggag caggaaactc aggagaaggg cagtgggcaa
     3661 gaggaggaga aaggggtggt cactaatggc ttagatgtga gccctgcgga ggaaaagaaa
     3721 ggggaggata gaagtgatga caaagtggtg gtgaccaaga aggtagaaaa aatcaccagc
     3781 gagggaggcg atggtgctac caaatacatc accaaatctg ttactgtcac tcaaaaggtt
     3841 gaagagcatg aggagacctt tgaggagaag ctggtgtcaa ctaaaaaggt agaaaaggtc
     3901 acttcacatg ccatagtcaa ggaagtcacc cagggtgact aagatcggag tccattgcaa
     3961 aaggttaagc catacaacaa tttcaaaatg catgtgattg acagcttcaa aacagaatgg
     4021 gttctcccat gagggctcca gacattgtat tttcctttgt gcaatatgag ggaactgcat
     4081 gcaagctcag ggtgcccccc tcctcagtcc ttgggggaat tgggatccac cggatctaga
     4141 taactgatca taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc
     4201 ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt
     4261 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     4321 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaacgcgta
     4381 aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt
     4441 ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat
     4501 agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa
     4561 cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta
     4621 atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc
     4681 ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc
     4741 gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac
     4801 acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt
     4861 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     4921 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag
     4981 aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     5041 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     5101 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     5161 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     5221 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     5281 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac
     5341 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
     5401 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
     5461 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
     5521 cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg
     5581 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
     5641 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
     5701 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
     5761 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
     5821 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
     5881 caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
     5941 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
     6001 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     6061 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     6121 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc
     6181 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
     6241 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     6301 ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca
     6361 ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt
     6421 gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca
     6481 ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca
     6541 agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc
     6601 tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     6661 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     6721 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     6781 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     6841 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     6901 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     6961 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     7021 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     7081 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     7141 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     7201 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     7261 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     7321 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     7381 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     7441 gataaccgta ttaccgccat gcat
//
