LOCUS       sequence-273922-        6540 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  M1 SpastinHFDD expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6540)
  AUTHORS   Chang CL, Weigel AV, Ioannou MS, Pasolli HA, Xu CS, Peale DR, 
            Shtengel G, Freeman M, Hess HF, Blackstone C, Lippincott-Schwartz J
  TITLE     Spastin tethers lipid droplets to peroxisomes and directs fatty acid
            trafficking through ESCRT-III.
  JOURNAL   J Cell Biol. 2019 Aug 5;218(8):2583-2599. doi: 
            10.1083/jcb.201902061. Epub 2019 Jun 21.
  PUBMED    31227594
REFERENCE   2  (bases 1 to 6540)
  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: 
            "10.1083/jcb.201902061"; journalName: "J Cell Biol"; date: 
            "2019-08-5- 5"; volume: "218"; issue: "8"; pages: "2583-2599"
COMMENT     Sequence Label: mApple-M1 SpastinHFDD
FEATURES             Location/Qualifiers
     source          1..6540
                     /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"
     regulatory      547..556
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             553..1260
                     /codon_start=1
                     /product="photostable monomeric derivative of DsRed (Shaner
                     et al., 2008)"
                     /label=mApple
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEA
                     FQTAKLKVTKGGPLPFAWDILSPQFMYGSKVYIKHPADIPDYFKLSFPEGFRWERVMNF
                     EDGGIIHVNQDSSLQDGVFIYKVKLRGTNFPSDGPVMQKKTMGWEASEERMYPEDGALK
                     SEIKKRLKLKDGGHYAAEVKTTYKAKKPVQLPGAYIVDIKLDIVSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     966..985
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    3268..3389
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3305..3324)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3359..3378
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3396..3851)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3533..3554)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3745..3764
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3878..3982
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3984..4341
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3984..4175
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4007..4027)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4192..4327
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4376..5170
                     /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             4376..5170
                     /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(4430..4449)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5040..5059
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5358..5377)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5402..5449
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5778..6366
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6267..6286
                     /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 gctagcgcta
      541 ccggtcgcca ccatggtgag caagggcgag gagaataaca tggccatcat caaggagttc
      601 atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc
      661 gagggcgagg gccgccccta cgaggccttt cagaccgcta agctgaaggt gaccaagggt
      721 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggtc
      781 tacattaagc acccagccga catccccgac tacttcaagc tgtccttccc cgagggcttc
      841 aggtgggagc gcgtgatgaa cttcgaggac ggcggcatta ttcacgttaa ccaggactcc
      901 tccctgcagg acggcgtgtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc
      961 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctccgagga gcggatgtac
     1021 cccgaggacg gcgccctgaa gagcgagatc aagaagaggc tgaagctgaa ggacggcggc
     1081 cactacgccg ccgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc
     1141 gcctacatcg tcgacatcaa gttggacatc gtgtcccaca acgaggacta caccatcgtg
     1201 gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag
     1261 tccggactca gatctcgagc tcaagcttcg aattctccgg gtggacgagg gaagaagaaa
     1321 ggctccggcg gcgccagcaa cccggtgcct cccaggcctc cgcccccttg cctggccccc
     1381 gcccctcccg ccgccgggcc ggcccctccg cccgagtcgc cgcataagcg gaacctgtac
     1441 tatttctcct acccgctgtt tgtaggcttc gcgctgctgc gtttggtcgc cttccacctg
     1501 gggctcctct tcgtgtggct ctgccagcgc ttctcccgcg ccctcgcggc agccaagagg
     1561 agctccgggg ccgcgccagc acctgcctcg gcctcggccc cggcgccggt gccgggcggc
     1621 gaggccgagc gcgtccgagt cttcgacaaa caggccgacg agtacatctc cattgccctg
     1681 cgcatcgatg aggatgagaa agcaggacag aaggagcaag ctgtggaatg gtataagaaa
     1741 ggtattgaag aactggaaaa aggaatagct gttatagtta caggacaagg tgaacagtgt
     1801 gaaagagcta gacgccttca agctaaaatg atgactaatt tggttatggc caaggaccgc
     1861 ttacaacttc tagagaagat gcaaccagtt ttgccatttt ccaagtcaca aacggacgtc
     1921 tataatgaca gtactaactt ggcatgccgc aatggacatc tccagtcaga aagtggagct
     1981 gttccaaaaa gaaaagaccc cttaacacac actagtaatt cactgcctcg ttcaaaaaca
     2041 gttatgaaaa ctggatctgc aggcctttca ggccaccata gagcacctag ttacagtggt
     2101 ttatccatgg tttctggagt gaaacaggga tctggtcctg ctcctaccac tcataagggt
     2161 actccgaaaa caaataggac aaataaacct tctaccccta caactgctac tcgtaagaaa
     2221 aaagacttga agaattttag gaatgtggac agcaaccttg ctaaccttat aatgaatgaa
     2281 attgtggaca atggaacagc tgttaaattt gatgatatag ctggtcaaga cttggcaaaa
     2341 caagcattgc aagaaattgt tattcttcct tctctgaggc ctgagttgtt cacagggctt
     2401 agagctcctg ccagagggct gttactcttt ggtccacctg ggaatgggaa gacaatgctg
     2461 gctaaagcag tagctgcaga atcgaatgca accttcttta atataagtgc tgcaagttta
     2521 acttcaaaat acgtgggaga aggagagaaa ttggtgaggg ctctttttgc tgtggctcga
     2581 gaacttcaac cttctataat ttttatagat gaagttgata gccttttgtg tgaaagaaga
     2641 gaaggggagc acgatgctag tagacgccta aaaactgaat ttctaataga atttgatggt
     2701 gtacagtctg ctggagatga cagagtactt gtaatgggtg caactaatag gccacaagag
     2761 cttgatgagg ctgttctcag gcgtttcatc aaacgggtat atgtgtcttt accaaatgag
     2821 gagacaagac tacttttgct taaaaatctg ttatgtaaac aaggaagtcc attgacccaa
     2881 aaagaactag cacaacttgc tagaatgact gatggatact caggaagtga cctaacagct
     2941 ttggcaaaag atgcagcact gggtcctatc cgagaactaa aaccagaaca ggtgaagaat
     3001 atgtctgcca gtgagatgag aaatattcga ttatctgact tcactgaatc cttgaaaaaa
     3061 ataaaacgca gcgtcagccc tcaaacttta gaagcgtaca tacgttggaa caaggacttt
     3121 ggagatacca ctgtttaagg atccaccgga tctagataac tgatcataat cagccatacc
     3181 acatttgtag aggttttact tgctttaaaa aacctcccac acctccccct gaacctgaaa
     3241 cataaaatga atgcaattgt tgttgttaac ttgtttattg cagcttataa tggttacaaa
     3301 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     3361 ggtttgtcca aactcatcaa tgtatcttaa cgcgtaaatt gtaagcgtta atattttgtt
     3421 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     3481 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     3541 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     3601 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     3661 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     3721 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     3781 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     3841 acagggcgcg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     3901 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     3961 ataatattga aaaaggaaga gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag
     4021 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     4081 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     4141 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     4201 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     4261 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     4321 ggaggcctag gcttttgcaa agatcgatca agagacagga tgaggatcgt ttcgcatgat
     4381 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     4441 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     4501 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcaaga
     4561 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     4621 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     4681 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     4741 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     4801 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     4861 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgagcatgc ccgacggcga
     4921 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     4981 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     5041 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     5101 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     5161 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     5221 tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     5281 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     5341 cctaggggga ggctaactga aacacggaag gagacaatac cggaaggaac ccgcgctatg
     5401 acggcaataa aaagacagaa taaaacgcac ggtgttgggt cgtttgttca taaacgcggg
     5461 gttcggtccc agggctggca ctctgtcgat accccaccga gaccccattg gggccaatac
     5521 gcccgcgttt cttccttttc cccaccccac cccccaagtt cgggtgaagg cccagggctc
     5581 gcagccaacg tcggggcggc aggccctgcc atagcctcag gttactcata tatactttag
     5641 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     5701 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     5761 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     5821 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     5881 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     5941 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     6001 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     6061 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     6121 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     6181 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     6241 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     6301 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     6361 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     6421 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgccatgcat
     6481 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
//
