LOCUS       sequence-434127-        6317 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SAC1 with a C389S mutation in it with the TMD deleted and replaced 
            with the Fis1tail to replace the TMD and target the mitochondria 
            instead of the ER with a Blue Fluorophore.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6317)
  AUTHORS   Doyle CP, Rectenwald A, Timple L, Hammond GRV
  TITLE     Orthogonal Targeting of SAC1 to Mitochondria Implicates ORP2 as a 
            Major Player in PM PI4P Turnover.
  JOURNAL   Contact (Thousand Oaks). 2024 Feb 7;7:25152564241229272. doi: 
            10.1177/25152564241229272. eCollection 2024 Jan-Dec.
  PUBMED    38327560
REFERENCE   2  (bases 1 to 6317)
  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: "Contact 
            (Thousand Oaks)."; date: "2024-02-7"; volume: "7:25152564241229272. 
            doi"; pages: " 10.1177/25152564241229272. eCollection 2024 Jan-Dec"
COMMENT     Sequence Label: TagBFP2-C1-SACM1LdeltaTMD-C389S-Fis1tail
FEATURES             Location/Qualifiers
     source          1..6317
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             613..1314
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIK
                     VVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMAL
                     KLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARY
                     CDLPSKLGHKLN"
     polyA_signal    3105..3226
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3142..3161)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3196..3215
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3233..3688)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3370..3391)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3582..3601
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3715..3819
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3821..4178
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3821..4012
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3844..3864)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4029..4164
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4213..5007
                     /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             4213..5007
                     /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(4267..4286)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4877..4896
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5195..5214)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5239..5286
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5615..6203
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6104..6123
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag cgagctgatt aaggagaaca tgcacatgaa gctgtacatg
      661 gagggcaccg tggacaacca tcacttcaag tgcacatccg agggcgaagg caagccctac
      721 gagggcaccc agaccatgag aatcaaggtg gtcgagggcg gccctctccc cttcgccttc
      781 gacatcctgg ctactagctt cctctacggc agcaagacct tcatcaacca cacccagggc
      841 atccccgact tcttcaagca gtccttccct gagggcttca catgggagag agtcaccaca
      901 tacgaagacg ggggcgtgct gaccgctacc caggacacca gcctccagga cggctgcctc
      961 atctacaacg tcaagatcag aggggtgaac ttcacatcca acggccctgt gatgcagaag
     1021 aaaacactcg gctgggaggc cttcaccgag acgctgtacc ccgctgacgg cggcctggaa
     1081 ggcagaaacg acatggccct gaagctcgtg ggcgggagcc atctgatcgc aaacgccaag
     1141 accacatata gatccaagaa acccgctaag aacctcaaga tgcctggcgt ctactatgtg
     1201 gactacagac tggaaagaat caaggaggcc aacaacgaga cctacgtcga gcagcacgag
     1261 gtggcagtgg ccagatactg cgacctccct agcaaactgg ggcacaagct taattccgga
     1321 ctcagatctg agctcatggc gacggcggcc tacgagcagc tgaagctgca tatcacacct
     1381 gaaaaatttt atgtggaagc ttgtgatgat ggagcagatg acgtacttac cattgaccgt
     1441 gtgtccacag aggttaccct tgcagtcaag aaagatgttc ctccttcagc tgtcacaaga
     1501 ccaatatttg gtatactggg cacaatccat ctggtggcag gtaattatct tatagtcatt
     1561 accaaaaaga taaaagtagg tgaatttttc agtcatgtag tctggaaagc aacagatttt
     1621 gatgtccttt cttataagaa gacaatgttg cacttaactg atattcagtt acaagataat
     1681 aaaaccttcc tagcgatgct aaaccatgtc ttgaatgtgg atggatttta cttttcaaca
     1741 acatatgatt tgacccatac tttgcagcgg ctatccaaca ctagtcctga attccaagaa
     1801 atgagtctct tggaaagggc agatcagcgg tttgtatgga atggtcatct tctaagagaa
     1861 ctttctgcac agccagaggt tcatcggttt gcccttccag tgttacatgg ctttattacc
     1921 atgcattcat gttctattaa tggaaaatac tttgattgga ttctcatctc gaggaggagc
     1981 tgtttcagag ctggtgtgcg ctattatgta agaggaattg attcggaagg ccatgcagct
     2041 aactttgtag aaacagaaca aattgtgcac tacaatggga gcaaagcttc gtttgtacag
     2101 actcgaggat caatacctgt tttctggtcc caaagaccaa acctcaagta caaaccactg
     2161 ccacagatca gcaaagtagc aaatcacatg gacggtttcc aaaggcattt tgattcccaa
     2221 gtaattattt atggaaaaca agttataatc aatctgatta accagaaggg ctcggagaag
     2281 ccacttgagc agacatttgc aacaatggtg tcttccttgg gaagtggaat gatgagatac
     2341 attgcctttg acttccataa ggaatgtaaa aatatgagat gggatcgact aagtatttta
     2401 ttggatcagg tagcagaaat gcaagatgaa ttaagttatt ttctagtgga ctctgctggc
     2461 caggtggtgg caaaccagga aggcgtgttc cgaagcaata gcatggattg tctagataga
     2521 accaatgtga tccagagttt gttagctcgt cgttcacttc aggcccaact tcagagacta
     2581 ggagttttgc atgtgggaca aaagcttgaa gaacaagatg aatttgagaa gattttcaaa
     2641 aatgcctggg ctgacaacgc aaatgcttgt gccaagcaat atgcgggaac tggtgccttg
     2701 aagactgact ttaccagaac tggaaagaga actcatttgg gacttataat ggatggctgg
     2761 aactcaatga tacgatatta taagaacaac ttttccgatg gatttagaca agattccata
     2821 gacttatttc ttggaaacta ttcagtggat gaattagaat ctcatagtcc tttaagtgtt
     2881 ccaagggact ggaaattgtc gacggtaccg ggactcgtgg gcatggccat cgtgggaggc
     2941 atggccctgg gtgtggcggg actggccgga ctcatcggac ttgctgtgtc caagtccaaa
     3001 tcctgatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
     3061 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
     3121 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     3181 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta
     3241 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
     3301 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg
     3361 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
     3421 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt
     3481 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
     3541 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
     3601 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
     3661 gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga
     3721 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     3781 ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc
     3841 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     3901 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     3961 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     4021 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     4081 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     4141 agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga
     4201 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     4261 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     4321 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     4381 ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     4441 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     4501 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     4561 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     4621 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     4681 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     4741 agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     4801 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     4861 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     4921 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     4981 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     5041 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     5101 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     5161 tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg
     5221 aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt
     5281 ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac
     5341 cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg
     5401 gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt
     5461 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     5521 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     5581 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     5641 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     5701 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     5761 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     5821 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     5881 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     5941 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     6001 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     6061 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     6121 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     6181 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     6241 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     6301 gtattaccgc catgcat
//
