LOCUS       sequence-173634-        6400 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses L195V spastin M1 and M87 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6400)
  AUTHORS   Solowska JM, Garbern JY, Baas PW
  TITLE     Evaluation of loss of function as an explanation for SPG4-based 
            hereditary spastic paraplegia.
  JOURNAL   Hum Mol Genet. 2010 Jul 15;19(14):2767-79. doi: 10.1093/hmg/ddq177. 
            Epub 2010 Apr 29.
  PUBMED    20430936
REFERENCE   2  (bases 1 to 6400)
  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"; doi: 
            "10.1093/hmg/ddq177"; journalName: "Hum Mol Genet"; date: 
            "2010-07-15- 15"; volume: "19"; issue: "14"; pages: "2767-79"
COMMENT     Sequence Label: pCMV-Tag/L195V with 5'UTR
FEATURES             Location/Qualifiers
     source          1..6400
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        554..572
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2763..2779)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2764..2780)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2835..2854)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2836..2854)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    3128..3249
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3165..3184)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3219..3238
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3256..3711)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3393..3414)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3605..3624
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3738..3842
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3844..4201
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3844..4035
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3867..3887)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4052..4187
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4236..5030
                     /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             4236..5030
                     /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(4290..4309)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4900..4919
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5218..5237)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5262..5309
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5638..6226
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6127..6146
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgatt
      541 acgccaagct cgaaattaac cctcactaaa gggaacaaaa gctggaggac ccacgcgtcc
      601 gggcccgagc caccgactgc aggaggagaa ggggttgtgc tcctggccga ggaaggagaa
      661 aggggcgggg ccggcgggca gcgtgcggca gtgcggagct cctgagaccg gcgggcacac
      721 gggggtctgt ggcccccgcc gtagcagtgg ctgccgccgt cgcttggttc ccgtcggtct
      781 gcgggaggcg ggttatggcg gcggcggcag tgagagctgt gaatgaattc tccgggtgga
      841 cgagggaaga agaaaggctc cggcggcgcc agcaacccgg tgcctcccag gcctccgccc
      901 ccttgcctgg cccccgcccc tcccgccgcc gggccggccc ctccgcccga gtcgccgcat
      961 aagcggaacc tgtactattt ctcctacccg ctgtttgtag gcttcgcgct gctgcgtttg
     1021 gtcgccttcc acctggggct cctcttcgtg tggctctgcc agcgcttctc ccgcgccctc
     1081 atggcagcca agaggagctc cggggccgcg ccagcacctg cctcggcctc ggccccggcg
     1141 ccggtgccgg gcggcgaggc cgagcgcgtc cgagtcttcc acaaacaggc cttcgagtac
     1201 atctccattg ccctgcgcat cgatgaggat gagaaagcag gacagaagga gcaagctgtg
     1261 gaatggtata agaaaggtat tgaagaactg gaaaaaggaa tagctgttat agttacagga
     1321 caaggtgaac agtgtgaaag agctagacgc cttcaagcta aaatgatgac taatttggtt
     1381 atggccaagg accgcttaca acttgtagag aagatgcaac cagttttgcc attttccaag
     1441 tcacaaacgg acgtctataa tgacagtact aacttggcat gccgcaatgg acatctccag
     1501 tcagaaagtg gagctgttcc aaaaagaaaa gaccccttaa cacacactag taattcactg
     1561 cctcgttcaa aaacagttat gaaaactgga tctgcaggcc tttcaggcca ccatagagca
     1621 cctagttaca gtggtttatc catggtttct ggagtgaaac agggatctgg tcctgctcct
     1681 accactcata agggtactcc gaaaacaaat aggacaaata aaccttctac ccctacaact
     1741 gctactcgta agaaaaaaga cttgaagaat tttaggaatg tggacagcaa ccttgctaac
     1801 cttataatga atgaaattgt ggacaatgga acagctgtta aatttgatga tatagctggt
     1861 caagacttgg caaaacaagc attgcaagaa attgttattc ttccttctct gaggcctgag
     1921 ttgttcacag ggcttagagc tcctgccaga gggctgttac tctttggtcc acctgggaat
     1981 gggaagacaa tgctggctaa agcagtagct gcagaatcga atgcaacctt ctttaatata
     2041 agtgctgcaa gtttaacttc aaaatacgtg ggagaaggag agaaattggt gagggctctt
     2101 tttgctgtgg ctcgagaact tcaaccttct ataattttta tagatgaagt tgatagcctt
     2161 ttgtgtgaaa gaagagaagg ggagcacgat gctagtagac gcctaaaaac tgaatttcta
     2221 atagaatttg atggtgtaca gtctgctgga gatgacagag tacttgtaat gggtgcaact
     2281 aataggccac aagagcttga tgaggctgtt ctcaggcgtt tcatcaaacg ggtatatgtg
     2341 tctttaccaa atgaggagac aagactactt ttgcttaaaa atctgttatg taaacaagga
     2401 agtccattga cccaaaaaga actagcacaa cttgctagaa tgactgatgg atactcagga
     2461 agtgacctaa cagctttggc aaaagatgca gcactgggtc ctatccgaga actaaaacca
     2521 gaacaggtga agaatatgtc tgccagtgag atgagaaata ttcgattatc tgacttcact
     2581 gaatccttga aaaaaataaa acgcagcgtc agccctcaaa ctttagaagc gtacatacgt
     2641 tggaacaagg actttggaga taccactgtt taaggaaata cctttgtaaa cctgcagaac
     2701 attttactta aaagaggaaa cacaagatcc cccgggctgc aggaattcga tatcaagctt
     2761 atcgataccg tcgacctcga gggggggccc ggtaccttaa ttaattaagg taccaggtaa
     2821 gtgtacccaa ttcgccctat agtgagtcgt attacaattc actcgatcgc ccttcccaac
     2881 agttgcgcag cctgaatggc gaatggagat ccaattttta agtgtataat gtgttaaact
     2941 actgattcta attgtttgtg tattttagat tcacagtccc aaggctcatt tcaggcccct
     3001 cagtcctcac agtctgttca tgatcataat cagccatacc acatttgtag aggttttact
     3061 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     3121 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     3181 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     3241 tgtatcttaa cgcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     3301 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     3361 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     3421 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     3481 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     3541 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     3601 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     3661 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca
     3721 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     3781 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     3841 atcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     3901 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     3961 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     4021 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     4081 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     4141 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     4201 agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     4261 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     4321 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     4381 tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt
     4441 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     4501 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     4561 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     4621 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     4681 aagccggtct tgtcgatcag gatgatctgg acgaagaaca tcaggggctc gcgccagccg
     4741 aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg
     4801 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     4861 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     4921 ctgaagaact tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     4981 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     5041 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     5101 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     5161 cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga
     5221 aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa
     5281 taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca
     5341 ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc
     5401 cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc
     5461 aggccctgcc atagcctcag gttactcata tatactttag attgatttaa aacttcattt
     5521 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     5581 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     5641 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     5701 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     5761 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     5821 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     5881 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     5941 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     6001 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     6061 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     6121 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     6181 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     6241 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     6301 atcccctgat tctgtggata accgtattac cgccatgcat tagttattaa tagtaatcaa
     6361 ttacggggtc attagttcat agcccatata tggagttccg
//
