LOCUS       sequence-426351-        5370 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ER localization of low-affinity NFAST fused to Halo-ER .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5370)
  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 5370)
  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)-Halo-ER
FEATURES             Location/Qualifiers
     source          1..5370
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    337..458
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(374..393)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     428..447
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(465..920)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(602..623)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     814..833
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        947..1051
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1053..1410
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1053..1244
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1076..1096)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1261..1396
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             1445..2239
                     /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             1445..2239
                     /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(1499..1518)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2109..2128
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2427..2446)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2471..2518
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2847..3435
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3336..3355
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        3610..3913
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3914..4117
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4068..4088
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      4144..4153
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      4534..4543
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4540..5370,1..60)
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
ORIGIN
        1 aacccggacc tgatcggcag cgagatcgcg cgctggctgt cgacgctcga gatttccggc
       61 atgtacaagt ccggactcag atctcgagct caagcttcga tcaccaccgt ggagtccaac
      121 tcctcctggt ggaccaactg ggtgatcccc gccatctccg ccctggtggt ggccctgatg
      181 taccgcctgt acatggccga ggactaagga tccaccggat ctagataact gatcataatc
      241 agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg
      301 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
      361 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
      421 tctagttgtg gtttgtccaa actcatcaat gtatcttaac gcgtaaattg taagcgttaa
      481 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
      541 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
      601 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
      661 aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg
      721 gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg
      781 acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc
      841 tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa
      901 tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
      961 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     1021 aatgcttcaa taatattgaa aaaggaagag tcctgaggcg gaaagaacca gctgtggaat
     1081 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     1141 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     1201 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     1261 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     1321 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     1381 ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt
     1441 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     1501 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     1561 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     1621 actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     1681 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     1741 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     1801 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     1861 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     1921 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgagcatgcc
     1981 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     2041 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     2101 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     2161 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     2221 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     2281 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     2341 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     2401 ttcgcccacc ctagggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc
     2461 cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc gtttgttcat
     2521 aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg
     2581 ggccaatacg cccgcgtttc ttccttttcc ccaccccacc ccccaagttc gggtgaaggc
     2641 ccagggctcg cagccaacgt cggggcggca ggccctgcca tagcctcagg ttactcatat
     2701 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     2761 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     2821 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     2881 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     2941 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     3001 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     3061 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     3121 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     3181 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     3241 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     3301 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     3361 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     3421 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     3481 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     3541 gccatgcatt agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
     3601 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     3661 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     3721 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     3781 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     3841 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     3901 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
     3961 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
     4021 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
     4081 taggcgtgta cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatccg
     4141 ctagccacca tggagaccgt gagattcggc ggcgacgaca tcgagaacag cctggccaag
     4201 atggacgaca aggccctgga caagctggcc ttcggcgcca tccagctgga cggcaacggc
     4261 aagatcatcc actacaacgc cgccgagggc accatcaccg gcagagaccc caagaccgtg
     4321 atcggcaaga acttcttcac cgacgtggcc cccggcaccc agagcaagga gttccagggc
     4381 agattcaagg agggcgtgca gaagggcgac ctgaacacca tgttcgagtg gatgatcccc
     4441 accagcagag gccccaccaa ggtgaaggtg cacatgaaga aggccatgac ctccggagga
     4501 ggcggcagcg gcggaggggg atctctaccg gtcgccacca tggcagaaat cggtactggc
     4561 tttccattcg acccccatta tgtggaagtc ctgggcgagc gcatgcacta cgtcgatgtt
     4621 ggtccgcgcg atggcacccc tgtgctgttc ctgcacggta acccgacctc ctcctacgtg
     4681 tggcgcaaca tcatcccgca tgttgcaccg acccatcgct gcattgctcc agacctgatc
     4741 ggtatgggca aatccgacaa accagacctg ggttatttct tcgacgacca cgtccgcttc
     4801 atggatgcct tcatcgaagc cctgggtctg gaagaggtcg tcctggtcat tcacgactgg
     4861 ggctccgctc tgggtttcca ctgggccaag cgcaatccag agcgcgtcaa aggtattgca
     4921 tttatggagt tcatccgccc tatcccgacc tgggacgaat ggccagaatt tgcccgcgag
     4981 accttccagg ccttccgcac caccgacgtc ggccgcaagc tgatcatcga tcagaacgtt
     5041 tttatcgagg gtacgctgcc gatgggtgtc gtccgcccgc tgactgaagt cgagatggac
     5101 cattaccgcg agccgttcct gaatcctgtt gaccgcgagc cactgtggcg cttcccaaac
     5161 gagctgccaa tcgccggtga gccagcgaac atcgtcgcgc tggtcgaaga atacatggac
     5221 tggctgcacc agtcccctgt cccgaagctg ctgttctggg gcaccccagg cgttctgatc
     5281 ccaccggccg aagccgctcg cctggccaaa agcctgccta actgcaaggc tgtggacatc
     5341 ggcccgggtc tgaatctgct gcaagaagac
//
