LOCUS       Exported                4265 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Inducible expression of M87 N184X spastin in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pTRE/SP M87 N184X
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4265)
  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 4265)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4265
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    8..26
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    44..62
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    79..97
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    115..133
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    151..169
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    186..204
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    222..240
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     287..311
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      349..358
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     polyA_signal    2175..2256
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2212..2231)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2267..2284)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2438..3026)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2518..2537)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3197..4057)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3820..3839
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4058..4162)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4230..4248
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4265..314
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
ORIGIN
        1 agtttactcc ctatcagtga tagagaacgt atgtcgagtt tactccctat cagtgataga
       61 gaacgatgtc gagtttactc cctatcagtg atagagaacg tatgtcgagt ttactcccta
      121 tcagtgatag agaacgtatg tcgagtttac tccctatcag tgatagagaa cgtatgtcga
      181 gtttatccct atcagtgata gagaacgtat gtcgagttta ctccctatca gtgatagaga
      241 acgtatgtcg aggtaggcgt gtacggtggg aggcctatat aagcagagct cgtttagtga
      301 accgtcagat cgcctggaga attcgagctc ggtacccgct tctcccgcgc caccatggca
      361 gccaagagga gctccggggc cgcgccagca cctgcctcgg cctcggcccc ggcgccggtg
      421 ccgggcggcg aggccgagcg cgtccgagtc ttccacaaac aggccttcga gtacatctcc
      481 attgccctgc gcatcgatga ggatgagaaa gcaggacaga aggagcaagc tgtggaatgg
      541 tataagaaag gtattgaaga actggaaaaa ggaatagctg ttatagttac aggacaaggt
      601 gaacagtgtg aaagagctag acgccttcaa gctaaaatga tgacttaatt tggttatggc
      661 caaggaccgc ttacaacttc tagagaagat gcaaccagtt ttgccatttt ccaagtcaca
      721 aacggacgtc tataatgaca gtactaactt ggcatgccgc aatggacatc tccagtcaga
      781 aagtggagct gttccaaaaa gaaaagaccc cttaacacac actagtaatt cactgcctcg
      841 ttcaaaaaca gttatgaaaa ctggatctgc aggcctttca ggccaccata gagcacctag
      901 ttacagtggt ttatccatgg tttctggagt gaaacaggga tctggtcctg ctcctaccac
      961 tcataagggt actccgaaaa caaataggac aaataaacct tctaccccta caactgctac
     1021 tcgtaagaaa aaagacttga agaattttag gaatgtggac agcaaccttg ctaaccttat
     1081 aatgaatgaa attgtggaca atggaacagc tgttaaattt gatgatatag ctggtcaaga
     1141 cttggcaaaa caagcattgc aagaaattgt tattcttcct tctctgaggc ctgagttgtt
     1201 cacagggctt agagctcctg ccagagggct gttactcttt ggtccacctg ggaatgggaa
     1261 gacaatgctg gctaaagcag tagctgcaga atcgaatgca accttcttta atataagtgc
     1321 tgcaagttta acttcaaaat acgtgggaga aggagagaaa ttggtgaggg ctctttttgc
     1381 tgtggctcga gaacttcaac cttctataat ttttatagat gaagttgata gccttttgtg
     1441 tgaaagaaga gaaggggagc acgatgctag tagacgccta aaaactgaat ttctaataga
     1501 atttgatggt gtacagtctg ctggagatga cagagtactt gtaatgggtg caactaatag
     1561 gccacaagag cttgatgagg ctgttctcag gcgtttcatc aaacgggtat atgtgtcttt
     1621 accaaatgag gagacaagac tacttttgct taaaaatctg ttatgtaaac aaggaagtcc
     1681 attgacccaa aaagaactag cacaacttgc tagaatgact gatggatact caggaagtga
     1741 cctaacagct ttggcaaaag atgcagcact gggtcctatc cgagaactaa aaccagaaca
     1801 ggtgaagaat atgtctgcca gtgagatgag aaatattcga ttatctgact tcactgaatc
     1861 cttgaaaaaa ataaaacgca gcgtcagccc tcaaacttta gaagcgtaca tacgttggaa
     1921 caaggacttt ggagatacca ctgtttaagg aaataccttt gtaaacctgc agaacatttt
     1981 acttaaaaga ggaaacacaa gatcctctag tcagctgacg cgtgctagcg cggccgcatc
     2041 gataagcttg tcgacgatat ctctagagga tcataatcag ccataccaca tttgtagagg
     2101 ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg
     2161 caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     2221 tcacaaattt cacaaataaa gcattttttt cactgcctcg agcttcctcg ctcactgact
     2281 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     2341 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     2401 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     2461 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     2521 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     2581 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     2641 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     2701 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     2761 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     2821 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     2881 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     2941 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     3001 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     3061 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     3121 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     3181 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     3241 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     3301 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     3361 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     3421 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     3481 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     3541 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     3601 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     3661 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     3721 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     3781 tgcggcgacc gagttgctct tgcccggcgt caacacggga taataccgcg ccacatagca
     3841 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     3901 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     3961 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     4021 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     4081 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     4141 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     4201 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtcttc
     4261 actcg
//
