LOCUS       sequence-434125-        6299 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SAC1 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 6299)
  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 6299)
  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-Fis1tail
FEATURES             Location/Qualifiers
     source          1..6299
                     /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    3087..3208
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3124..3143)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3178..3197
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3215..3670)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3352..3373)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3564..3583
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3697..3801
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3803..4160
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3803..3994
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3826..3846)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4011..4146
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4195..4989
                     /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             4195..4989
                     /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(4249..4268)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4859..4878
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5177..5196)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5221..5268
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5597..6185
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6086..6105
                     /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 ctcagatcta tggcgacggc ggcctacgag cagctgaagc tgcatatcac acctgaaaaa
     1381 ttttatgtgg aagcttgtga tgatggagca gatgacgtac ttaccattga ccgtgtgtcc
     1441 acagaggtta cccttgcagt caagaaagat gttcctcctt cagctgtcac aagaccaata
     1501 tttggtatac tgggcacaat ccatctggtg gcaggtaatt atcttatagt cattaccaaa
     1561 aagataaaag taggtgaatt tttcagtcat gtagtctgga aagcaacaga ttttgatgtc
     1621 ctttcttata agaagacaat gttgcactta actgatattc agttacaaga taataaaacc
     1681 ttcctagcga tgctaaacca tgtcttgaat gtggatggat tttacttttc aacaacatat
     1741 gatttgaccc atactttgca gcggctatcc aacactagtc ctgaattcca agaaatgagt
     1801 ctcttggaaa gggcagatca gcggtttgta tggaatggtc atcttctaag agaactttct
     1861 gcacagccag aggttcatcg gtttgccctt ccagtgttac atggctttat taccatgcat
     1921 tcatgttcta ttaatggaaa atactttgat tggattctca tctcgaggag gagctgtttc
     1981 agagctggtg tgcgctatta tgtaagagga attgattcgg aaggccatgc agctaacttt
     2041 gtagaaacag aacaaattgt gcactacaat gggagcaaag cttcgtttgt acagactcga
     2101 ggatcaatac ctgttttctg gtcccaaaga ccaaacctca agtacaaacc actgccacag
     2161 atcagcaaag tagcaaatca catggacggt ttccaaaggc attttgattc ccaagtaatt
     2221 atttatggaa aacaagttat aatcaatctg attaaccaga agggctcgga gaagccactt
     2281 gagcagacat ttgcaacaat ggtgtcttcc ttgggaagtg gaatgatgag atacattgcc
     2341 tttgacttcc ataaggaatg taaaaatatg agatgggatc gactaagtat tttattggat
     2401 caggtagcag aaatgcaaga tgaattaagt tattttctag tggactctgc tggccaggtg
     2461 gtggcaaacc aggaaggcgt gttccgaagc aattgcatgg attgtctaga tagaaccaat
     2521 gtgatccaga gtttgttagc tcgtcgttca cttcaggccc aacttcagag actaggagtt
     2581 ttgcatgtgg gacaaaagct tgaagaacaa gatgaatttg agaagatttt caaaaatgcc
     2641 tgggctgaca acgcaaatgc ttgtgccaag caatatgcgg gaactggtgc cttgaagact
     2701 gactttacca gaactggaaa gagaactcat ttgggactta taatggatgg ctggaactca
     2761 atgatacgat attataagaa caacttttcc gatggattta gacaagattc catagactta
     2821 tttcttggaa actattcagt ggatgaatta gaatctcata gtcctttaag tgttccaagg
     2881 gactggaaat tgggactcgt gggcatggcc atcgtgggag gcatggccct gggtgtggcg
     2941 ggactggccg gactcatcgg acttgctgtg tccaagtcca aatcctgatc agccatacca
     3001 catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac
     3061 ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat
     3121 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     3181 gtttgtccaa actcatcaat gtatcttaac gcgtaaattg taagcgttaa tattttgtta
     3241 aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc
     3301 aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg
     3361 aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat
     3421 cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc
     3481 cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag
     3541 ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg
     3601 gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta
     3661 cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     3721 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     3781 taatattgaa aaaggaagag tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt
     3841 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3901 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     3961 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     4021 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     4081 cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg
     4141 gaggcctagg cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt
     4201 gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat
     4261 gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag
     4321 gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaagac
     4381 gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac
     4441 gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc
     4501 ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg
     4561 ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag
     4621 cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat
     4681 caggggctcg cgccagccga actgttcgcc aggctcaagg cgagcatgcc cgacggcgag
     4741 gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc
     4801 ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg
     4861 ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg
     4921 ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag
     4981 ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat
     5041 cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc
     5101 gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc
     5161 ctagggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc cgcgctatga
     5221 cggcaataaa aagacagaat aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg
     5281 ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg
     5341 cccgcgtttc ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg
     5401 cagccaacgt cggggcggca ggccctgcca tagcctcagg ttactcatat atactttaga
     5461 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     5521 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     5581 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     5641 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     5701 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     5761 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     5821 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     5881 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     5941 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     6001 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     6061 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     6121 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     6181 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     6241 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gccatgcat
//
