LOCUS       sequence-429341-        8182 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lipid droplet localization of 6xHp of Spastin fused to mApple.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8182)
  AUTHORS   Li X, Gamuyao R, Wu ML, Cho WJ, King SV, Petersen RA, Stabley DR, 
            Lindow C, Climer LK, Shirinifard A, Ferrara F, Throm RE, Robinson 
            CG, Zhou Y, Carisey AF, Tebo AG, Chang CL
  TITLE     A fluorogenic complementation tool kit for interrogating lipid 
            droplet-organelle interaction.
  JOURNAL   J Cell Biol. 2024 Sep 2;223(9):e202311126. doi: 
            10.1083/jcb.202311126. Epub 2024 Jul 1.
  PUBMED    38949658
REFERENCE   2  (bases 1 to 8182)
  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: "J Cell 
            Biol."; date: "2024-09-2"; volume: "223(9):e202311126. doi"; pages: 
            " 10.1083/jcb.202311126. Epub 2024 Jul 1"
COMMENT     Sequence Label: Lenti-mApple-6xHp
FEATURES             Location/Qualifiers
     source          1..8182
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(201..908)
                     /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     complement(476..495)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     regulatory      905..914
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(961..1164)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     enhancer        1165..1468
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     CDS             complement(1725..1769)
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    1954..2187
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    2547..2664
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_feature    3156..3281
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             3328..3508
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(3523..3726)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     enhancer        3727..4106
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     CDS             join(4245..4313,4314..5105)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4463..4482)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5338..5926
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5827..5846
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6080..6097
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    6165..6299
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6189..6208)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6243..6262
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(6305..6500)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      6319..6454
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     protein_bind    6559..6580
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6595..6600,6601..6618,6619..6625)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6633..6649
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     6638..6660
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6657..6673
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6657..6673
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7016..7035
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    7034..7089
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     7135..7154
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
ORIGIN
        1 gattcactat tagccgaacg tttcgcagcc atcagcgctc tagaaaagcg ttgacacaac
       61 caaacaaaca gtaatccaag atgaaaagca accaatcgaa gtagagcaaa cccgacaaaa
      121 agggggtaag agaagtagta caggttgcgt ttgtgcgggg attcggaatt cgatgcttga
      181 gctcgagatc tgagtccgga cttgtacagc tcgtccatgc cgccggtgga gtggcggccc
      241 tcggcgcgtt cgtactgttc cacgatggtg tagtcctcgt tgtgggacac gatgtccaac
      301 ttgatgtcga cgatgtaggc gccgggcagc tgcacgggct tcttggcctt gtaggtggtc
      361 ttgacctcgg cggcgtagtg gccgccgtcc ttcagcttca gcctcttctt gatctcgctc
      421 ttcagggcgc cgtcctcggg gtacatccgc tcctcggagg cctcccagcc catggtcttc
      481 ttctgcatta cggggccgtc ggaggggaag ttggtgccgc gcagcttcac cttgtagatg
      541 aacacgccgt cctgcaggga ggagtcctgg ttaacgtgaa taatgccgcc gtcctcgaag
      601 ttcatcacgc gctcccacct gaagccctcg gggaaggaca gcttgaagta gtcggggatg
      661 tcggctgggt gcttaatgta gaccttggag ccgtacatga actgagggga caggatgtcc
      721 caggcgaagg gcagggggcc acccttggtc accttcagct tagcggtctg aaaggcctcg
      781 taggggcggc cctcgccctc gccctcgatc tcgaactcgt ggccgttcac ggagccctcc
      841 atgtgcacct tgaagcgcat gaactccttg atgatggcca tgttattctc ctcgcccttg
      901 ctcaccatgg tggcgaccgg tagctcgagg gttcactaaa cgagcccggt tcactaaacg
      961 agctctgctt atatagacct cccaccgtac acgcctaccg cccatttgcg tcaatggggc
     1021 ggagttgtta cgacattttg gaaagtcccg ttgattttgg tgccaaaaca aactcccatt
     1081 gacgtcaatg gggtggagac ttggaaatcc ccgtgagtca aaccgctatc cacgcccatt
     1141 gatgtactgc caaaaccgca tcaccatggt aatagcgatg actaatacgt agatgtactg
     1201 ccaagtagga aagtcccata aggtcatgta ctgggcataa tgccaggcgg gccatttacc
     1261 gtcattgacg tcaatagggg gcgtacttgg catatgatac acttgatgta ctgccaagtg
     1321 ggcagtttac cgtaaatact ccacccattg acgtcaatgg aaagtcccta ttggcgttac
     1381 tatgggaaca tacgtcatta ttgacgtcaa tgggcggggg tcgttgggcg gtcagccagg
     1441 cgggccattt accgtaagtt atgtaacgcg gaactccata tatgggctat gaactaatga
     1501 ccccgtaatt gattactatt aataactagc acgcgttcga agcttgagct cggtcccctc
     1561 ggggttggga ggtgggtctg aaacgataat ggtgaatatc cctgcctaac tctattcact
     1621 atagaaagta cagcaaaaac tattcttaaa cctaccaagc ctcctactat cattatgaat
     1681 aattttatat accacagcca atttgttatg ttaaaccaat tccacaaact tgcccattta
     1741 tctaattcca ataattcttg ttcattcttt tcttgctggt tttgcgattc ttcaattaag
     1801 gagtgtatta agcttgtgta attgttaatt tctctgtccc actccatcca ggtcgtgtga
     1861 ttccaaatct gttccagaga tttattactc caactagcat tccaaggcac agcagtggtg
     1921 caaatgagtt ttccagagca accccaaatc cccaggagct gttgatcctt taggtatctt
     1981 tccacagcca ggattcttgc ctggagctgc ttgatgcccc agactgtgag ttgcaacaga
     2041 tgctgttgcg cctcaatagc cctcagcaaa ttgttctgct gctgcactat accagacaat
     2101 aattgtctgg cctgtaccgt cagcgtcatt gacgctgcgc ccatagtgct tcctgctgct
     2161 cccaagaacc caaggaacaa agctcctatt cccactgctc ttttttctct ctgcaccact
     2221 cttctctttg ccttggtggg tgctactcct aatggttcaa tttttactac tttatattta
     2281 tataattcac ttctccaatt gtccctcata tctcctcctt atggtgtttt actaaacttt
     2341 tccatgttct aatcctcatc ctgtctactt gccacacaat catcacctgc catctgtttt
     2401 ccataatccc taatgatctt tgcttttctt cttggcacta cttttatgtc actattatct
     2461 tgtattacta ctgccccttc acctttccag aggagctttg ctggtccttt ccaaagtgga
     2521 tttctgctgt ccctgtaata aacccgaaaa ttttgaattt ttgtaatttg tttttgtaat
     2581 tctttagttt gtatgtctgt tgctattatg tctactattc tttcccctgc actgtacccc
     2641 ccaatccccc cttttctttt aaaattgtgg atgaatactg ccatttgtac tgctgtctta
     2701 agatgttcag cctgatctct tacctgtcct ataattttct ttaattcttt attcatagat
     2761 tctactactc cttgactttg gggattgtag ggaatttcca ggtctgaaga tcctgctgct
     2821 tgctgtgctt ttttcttact tttgttttgc tcttcctcta tcttgtctaa agcttccttg
     2881 gtgtctttta tctctatcct ttgatgcaca caatagaggg ttgctactgt attatataat
     2941 gatctaagtt cttctgatcc tgtctgaagg gatggttgta gctgtcccag tatttgtcta
     3001 cagccttctg atgtttctaa caggccagta ttaactgcga atcgttctag ctccctgctt
     3061 gcccatacta tatgttttaa tttatatttt ttctttcccc ctggccttaa ccgaattttt
     3121 tcccatcgcg atctaattct cccccgctta atactgacgc tctcgcaccc atctctctcc
     3181 ttctagcctc cgctagtcaa aatttttggc gtactcacca gtcgccgccc ctcgcctctt
     3241 gccgtgcgcg cttcagcaag ccgagtcctg cgtcgagaga gctcctctgg tttccctttc
     3301 gctttcaagt ccctgttcgg gcgccactgc tagagatttt ccacactgac taaaagggtc
     3361 tgagggatct ctagttacca gagtcacaca acagacgggc acacactact tgaagcactc
     3421 aaggcaagct ttattgaggc ttaagcagtg ggttccctag ttagccagag agctcccagg
     3481 ctcagatctg gtctaaccag agagaccccg gttcactaaa cgagctctgc ttatatagac
     3541 ctcccaccgt acacgcctac cgcccatttg cgtcaatggg gcggagttgt tacgacattt
     3601 tggaaagtcc cgttgatttt ggtgccaaaa caaactccca ttgacgtcaa tggggtggag
     3661 acttggaaat ccccgtgagt caaaccgcta tccacgccca ttgatgtact gccaaaaccg
     3721 catcaccatg gtaatagcga tgactaatac gtagatgtac tgccaagtag gaaagtccca
     3781 taaggtcatg tactgggcat aatgccaggc gggccattta ccgtcattga cgtcaatagg
     3841 gggcgtactt ggcatatgat acacttgatg tactgccaag tgggcagttt accgtaaata
     3901 ctccacccat tgacgtcaat ggaaagtccc tattggcgtt actatgggaa catacgtcat
     3961 tattgacgtc aatgggcggg ggtcgttggg cggtcagcca ggcgggccat ttaccgtaag
     4021 ttatgtaacg cggaactcca tatatgggct atgaactaat gaccccgtaa ttgattacta
     4081 ttaataacta gtcaataatc aatgtcgacc caggtggcac ttttcgggga aatgtgcgcg
     4141 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     4201 aaccctgata aatgcttcaa taataattct agacgtctcc agcaatgagt attcaacatt
     4261 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     4321 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     4381 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     4441 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     4501 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     4561 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     4621 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     4681 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     4741 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     4801 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     4861 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     4921 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     4981 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     5041 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     5101 ggtaaagcac gagacgtcta gaatgatttt tatcaaaatc attaagttaa ggtagataca
     5161 catcttgtca tatgatcaaa tggtttcgcc aaaaatcaat aatcagacaa caagatgtgc
     5221 gaactcgata ttttacacga ctctctttac caattctacc accatgacca aaatccctta
     5281 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     5341 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     5401 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     5461 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     5521 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     5581 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     5641 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     5701 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     5761 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     5821 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     5881 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     5941 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     6001 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     6061 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     6121 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agcggatcca gacatgataa
     6181 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt
     6241 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta
     6301 gctttttgca aaagcctagg cctccaaaaa agcctcctca ctacttctgg aatagctcag
     6361 aggccgaggc ggcctcggcc tctgcataaa taaaaaaaat tagtcagcca tggggcggag
     6421 aatgggcgga actgggcgga gttaggggcg ggatgggcgg agttaggggc gggactatgg
     6481 ttgctgacta attgagatgc ggatccgctg gcacgacagg tttcccgact ggaaagcggg
     6541 cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca
     6601 ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg
     6661 aaacagctat gaccatgatt acgccaagct tgggctgcag gtcgagggat ctccataaga
     6721 gaagagggac agctatgact gggagtagtc aggagaggag gaaaaatctg gctagtaaaa
     6781 catgtaagga aaattttagg gatgttaaag aaaaaaataa cacaaaacaa aatataaaaa
     6841 aaatctaacc tcaagtcaag gcttttctat ggaataagga atggacagca gggggctgtt
     6901 tcatatactg atgacctctt tatagccaac ctttgttcat ggcagccagc atatgggcat
     6961 atgttgccaa actctaaacc aaatactcat tctgatgttt taaatgattt gccctcccat
     7021 atgtccttcc gagtgagaga cacaaaaaat tccaacacac tattgcaatg aaaataaatt
     7081 tcctttatta gccagaagtc agatgctcaa ggggcttcat gatgtcccca taatttttgg
     7141 cagagggaaa aagatctgct agcacgacaa ggcaagcttt attgaggctt aagcagtggg
     7201 ttccctagtt agccagagag ctcccaggct cagatctggt ctaaccagag agacccagta
     7261 caggcaaaaa gcagatcttg tcttctttgg gagtgaatta gcccttccag tccccccttt
     7321 tcttttaaaa agtggctaag atctacagct gccttgtaag tcattggtct taaaggtacc
     7381 tagtgcggcc gcggatcctt aggagcgctt ggctgccatt agcgcccggg agaatctctg
     7441 gcaaagccat acgaacagca agccgaggtg gaatgcaacc agcctgagta acgcgaaacc
     7501 gacgaagagc gggtaactaa agtaatagag gttccgcttg tgaggagatt cagaatttgc
     7561 gcttctcttc gcagccatta aggctcgtga aaatcgctgg cacaaccaga cgaataataa
     7621 tcccagatga aatgctacaa gacgtaatag tgcgaaccct acgaagagag gatatgagaa
     7681 atagtacaga ttccgcttat gaggtgattc gctatttgcg gaacgcttag ctgccatcaa
     7741 ggctcgcgag aagcgctggc agagccagac gaatagtagg cctagatgga aggccactag
     7801 ccggagtaga gcaaagccta caaaaagagg gtaagagaag tagtataagt tccgcttatg
     7861 cggggattcg ctattggcac tcctcttagc cgccattaaa gctcgcgaaa atctttggca
     7921 aagccacacg aacagaagac caagatgaaa cgcaactagc cgaaggagag cgaagccaac
     7981 gaacaacggg taagagaagt aatagaggtt gcgtttatga ggcgattcgc tatttgccga
     8041 cctctttgct gccattaacg ctcgcgagaa ccgttgacat aaccagacaa acaacaaacc
     8101 tagatgaaac gcgaccagcc gaagtagcgc gaaacccacg aacaagggat agctgaagta
     8161 atataagttc ctcttatggg gc
//
