LOCUS       sequence-181056-        5924 bp    DNA     circular SYN 15-APR-2025
DEFINITION  expression of M87 S245X spastin in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5924)
  AUTHORS   Solowska JM, Rao AN, Baas PW
  TITLE     Studies on truncating mutations of SPAST associated with Hereditary 
            Spastic Paraplegia indicate greater accumulation and toxicity of the
            M1 isoform of spastin.
  JOURNAL   Mol Biol Cell. 2017 May 11. pii: mbc.E17-01-0047. doi: 
            10.1091/mbc.E17-01-0047.
  PUBMED    28495799
REFERENCE   2  (bases 1 to 5924)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Biol 
            Cell. 2017 May 11. pii: mbc.E17-01-0047. doi: 
            10.1091/mbc.E17-01-0047."
COMMENT     Sequence Label: pCMV-Tag/M87 S245X
FEATURES             Location/Qualifiers
     source          1..5924
                     /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"
     regulatory      599..608
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(2287..2303)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2288..2304)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2359..2378)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2360..2378)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    2652..2773
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2689..2708)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2743..2762
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2780..3235)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2917..2938)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3129..3148
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3262..3366
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3368..3725
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3368..3559
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3391..3411)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3576..3711
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3760..4554
                     /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             3760..4554
                     /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(3814..3833)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4424..4443
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4742..4761)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4786..4833
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5162..5750
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5651..5670
                     /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 gctggaggct tctcccgcgc
      601 caccatggca gccaagagga gctccggggc cgcgccagca cctgcctcgg cctcggcccc
      661 ggcgccggtg ccgggcggcg aggccgagcg cgtccgagtc ttccacaaac aggccttcga
      721 gtacatctcc attgccctgc gcatcgatga ggatgagaaa gcaggacaga aggagcaagc
      781 tgtggaatgg tataagaaag gtattgaaga actggaaaaa ggaatagctg ttatagttac
      841 aggacaaggt gaacagtgtg aaagagctag acgccttcaa gctaaaatga tgactaattt
      901 ggttatggcc aaggaccgct tacaacttct agagaagatg caaccagttt tgccattttc
      961 caagtcacaa acggacgtct ataatgacag tactaacttg gcatgccgca atggacatct
     1021 ccagtcagaa agtggagctg ttccaaaaag aaaagacccc ttaacacaca ctagtaattg
     1081 actgcctcgt tcaaaaacag ttatgaaaac tggatctgca ggcctttcag gccaccatag
     1141 agcacctagt tacagtggtt tatccatggt ttctggagtg aaacagggat ctggtcctgc
     1201 tcctaccact cataagggta ctccgaaaac aaataggaca aataaacctt ctacccctac
     1261 aactgctact cgtaagaaaa aagacttgaa gaattttagg aatgtggaca gcaaccttgc
     1321 taaccttata atgaatgaaa ttgtggacaa tggaacagct gttaaatttg atgatatagc
     1381 tggtcaagac ttggcaaaac aagcattgca agaaattgtt attcttcctt ctctgaggcc
     1441 tgagttgttc acagggctta gagctcctgc cagagggctg ttactctttg gtccacctgg
     1501 gaatgggaag acaatgctgg ctaaagcagt agctgcagaa tcgaatgcaa ccttctttaa
     1561 tataagtgct gcaagtttaa cttcaaaata cgtgggagaa ggagagaaat tggtgagggc
     1621 tctttttgct gtggctcgag aacttcaacc ttctataatt tttatagatg aagttgatag
     1681 ccttttgtgt gaaagaagag aaggggagca cgatgctagt agacgcctaa aaactgaatt
     1741 tctaatagaa tttgatggtg tacagtctgc tggagatgac agagtacttg taatgggtgc
     1801 aactaatagg ccacaagagc ttgatgaggc tgttctcagg cgtttcatca aacgggtata
     1861 tgtgtcttta ccaaatgagg agacaagact acttttgctt aaaaatctgt tatgtaaaca
     1921 aggaagtcca ttgacccaaa aagaactagc acaacttgct agaatgactg atggatactc
     1981 aggaagtgac ctaacagctt tggcaaaaga tgcagcactg ggtcctatcc gagaactaaa
     2041 accagaacag gtgaagaata tgtctgccag tgagatgaga aatattcgat tatctgactt
     2101 cactgaatcc ttgaaaaaaa taaaacgcag cgtcagccct caaactttag aagcgtacat
     2161 acgttggaac aaggactttg gagataccac tgtttaagga aatacctttg taaacctgca
     2221 gaacatttta cttaaaagag gaaacacaag atcccccggg ctgcaggaat tcgatatcaa
     2281 gcttatcgat accgtcgacc tcgagggggg gcccggtacc ttaattaatt aaggtaccag
     2341 gtaagtgtac ccaattcgcc ctatagtgag tcgtattaca attcactcga tcgcccttcc
     2401 caacagttgc gcagcctgaa tggcgaatgg agatccaatt tttaagtgta taatgtgtta
     2461 aactactgat tctaattgtt tgtgtatttt agattcacag tcccaaggct catttcaggc
     2521 ccctcagtcc tcacagtctg ttcatgatca taatcagcca taccacattt gtagaggttt
     2581 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
     2641 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     2701 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     2761 tcaatgtatc ttaacgcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     2821 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     2881 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     2941 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     3001 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     3061 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     3121 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     3181 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt
     3241 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     3301 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     3361 aagaatcctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt
     3421 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     3481 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     3541 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     3601 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     3661 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     3721 gcaaagatcg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     3781 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     3841 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     3901 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta
     3961 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     4021 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     4081 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     4141 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     4201 atggaagccg gtcttgtcga tcaggatgat ctggacgaag aacatcaggg gctcgcgcca
     4261 gccgaactgt tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc
     4321 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     4381 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     4441 attgctgaag aacttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     4501 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     4561 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     4621 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     4681 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa
     4741 ctgaaacacg gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac
     4801 agaataaaac gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct
     4861 ggcactctgt cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct
     4921 tttccccacc ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg
     4981 cggcaggccc tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc
     5041 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     5101 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     5161 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     5221 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     5281 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     5341 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     5401 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     5461 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     5521 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     5581 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     5641 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     5701 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     5761 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     5821 cgttatcccc tgattctgtg gataaccgta ttaccgccat gcattagtta ttaatagtaa
     5881 tcaattacgg ggtcattagt tcatagccca tatatggagt tccg
//
