LOCUS       sequence-426326-        6024 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lipid droplet localization of low-affinity NFAST fused to 
            mApple-6xHp.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6024)
  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 6024)
  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: NFAST(low)-mApple-6xHp
FEATURES             Location/Qualifiers
     source          1..6024
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        16..319
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        320..523
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     474..494
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      550..559
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      940..949
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             946..1653
                     /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     1359..1378
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2767..2888
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2804..2823)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2858..2877
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2895..3350)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3032..3053)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3244..3263
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3377..3481
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3483..3840
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3483..3674
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3506..3526)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3691..3826
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3875..4669
                     /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             3875..4669
                     /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(3929..3948)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4539..4558
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4857..4876)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4901..4948
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5277..5865
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5766..5785
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
       61 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      121 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      181 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      241 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      301 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
      361 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
      421 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
      481 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca
      541 gatccgctag ccaccatgga gaccgtgaga ttcggcggcg acgacatcga gaacagcctg
      601 gccaagatgg acgacaaggc cctggacaag ctggccttcg gcgccatcca gctggacggc
      661 aacggcaaga tcatccacta caacgccgcc gagggcacca tcaccggcag agaccccaag
      721 accgtgatcg gcaagaactt cttcaccgac gtggcccccg gcacccagag caaggagttc
      781 cagggcagat tcaaggaggg cgtgcagaag ggcgacctga acaccatgtt cgagtggatg
      841 atccccacca gcagaggccc caccaaggtg aaggtgcaca tgaagaaggc catgacctcc
      901 ggaggaggcg gcagcggcgg agggggatct ctaccggtcg ccaccatggt gagcaagggc
      961 gaggagaata acatggccat catcaaggag ttcatgcgct tcaaggtgca catggagggc
     1021 tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgaggcc
     1081 tttcagaccg ctaagctgaa ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc
     1141 ctgtcccctc agttcatgta cggctccaag gtctacatta agcacccagc cgacatcccc
     1201 gactacttca agctgtcctt ccccgagggc ttcaggtggg agcgcgtgat gaacttcgag
     1261 gacggcggca ttattcacgt taaccaggac tcctccctgc aggacggcgt gttcatctac
     1321 aaggtgaagc tgcgcggcac caacttcccc tccgacggcc ccgtaatgca gaagaagacc
     1381 atgggctggg aggcctccga ggagcggatg taccccgagg acggcgccct gaagagcgag
     1441 atcaagaaga ggctgaagct gaaggacggc ggccactacg ccgccgaggt caagaccacc
     1501 tacaaggcca agaagcccgt gcagctgccc ggcgcctaca tcgtcgacat caagttggac
     1561 atcgtgtccc acaacgagga ctacaccatc gtggaacagt acgaacgcgc cgagggccgc
     1621 cactccaccg gcggcatgga cgagctgtac aagtccggac tcagatctcg agctcaagct
     1681 tcgaattctg agtcgccgca taagcggaac ctgtactatt tctcctaccc gctgtttgta
     1741 ggcttcgcgc tgctgcgttt ggtcgccttc cacctggggc tcctcttcgt gtggctctgc
     1801 cagcgcttct cccgcgccct catggcagcc aagaggagcg ccaattctga gtcgccgcat
     1861 aagcggaacc tgtactattt ctcctacccg ctgtttgtag gcttcgcgct gctgcgtttg
     1921 gtcgccttcc acctggggct cctcttcgtg tggctctgcc agcgcttctc ccgcgccctc
     1981 atggcagcca agaggagcgc caattctgag tcgccgcata agcggaacct gtactatttc
     2041 tcctacccgc tgtttgtagg cttcgcgctg ctgcgtttgg tcgccttcca cctggggctc
     2101 ctcttcgtgt ggctctgcca gcgcttctcc cgcgccctca tggcagccaa gaggagcgcc
     2161 aattctgagt cgccgcataa gcggaacctg tactatttct cctacccgct gtttgtaggc
     2221 ttcgcgctgc tgcgtttggt cgccttccac ctggggctcc tcttcgtgtg gctctgccag
     2281 cgcttctccc gcgccctcat ggcagccaag aggagcgcca attctgagtc gccgcataag
     2341 cggaacctgt actatttctc ctacccgctg tttgtaggct tcgcgctgct gcgtttggtc
     2401 gccttccacc tggggctcct cttcgtgtgg ctctgccagc gcttctcccg cgccctcatg
     2461 gcagccaaga ggagcgccaa ttctgagtcg ccgcataagc ggaacctgta ctatttctcc
     2521 tacccgctgt ttgtaggctt cgcgctgctg cgtttggtcg ccttccacct ggggctcctc
     2581 ttcgtgtggc tctgccagcg cttctcccgc gccctcatgg cagccaagag gagctaagga
     2641 tccaccggat ctagataact gatcataatc agccatacca catttgtaga ggttttactt
     2701 gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt
     2761 gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     2821 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     2881 gtatcttaac gcgtaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg
     2941 ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa
     3001 agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa
     3061 gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg
     3121 tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa
     3181 ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa
     3241 ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct
     3301 gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac
     3361 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     3421 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     3481 tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
     3541 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
     3601 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     3661 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     3721 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg
     3781 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     3841 gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc
     3901 aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat
     3961 cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt
     4021 caagaccgac ctgtccggtg ccctgaatga actgcaagac gaggcagcgc ggctatcgtg
     4081 gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag
     4141 ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc
     4201 tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc
     4261 tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga
     4321 agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga
     4381 actgttcgcc aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg
     4441 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
     4501 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
     4561 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
     4621 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
     4681 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
     4741 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     4801 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ctagggggag gctaactgaa
     4861 acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa aagacagaat
     4921 aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac
     4981 tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc
     5041 ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca
     5101 ggccctgcca tagcctcagg ttactcatat atactttaga ttgatttaaa acttcatttt
     5161 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     5221 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     5281 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     5341 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     5401 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     5461 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     5521 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     5581 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     5641 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     5701 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     5761 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     5821 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     5881 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     5941 tcccctgatt ctgtggataa ccgtattacc gccatgcatt agttattaat agtaatcaat
     6001 tacggggtca ttagttcata gccc
//
