LOCUS       sequence-468028-        5787 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of CFAST10 on the OMM membrane after a long linker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5787)
  AUTHORS   Garcia Casas P, Rossini M, Pavenius L, Saeed M, Arnst N, Sonda S, 
            Fernandes T, D'Arsie I, Bruzzone M, Berno V, Raimondi A, Sassano ML,
            Naia L, Barbieri E, Sigismund S, Agostinis P, Sturlese M, Niemeyer 
            BA, Brismar H, Ankarcrona M, Gautier A, Pizzo P, Filadi R
  TITLE     Simultaneous detection of membrane contact dynamics and associated 
            Ca(2+) signals by reversible chemogenetic reporters.
  JOURNAL   Nat Commun. 2024 Nov 12;15(1):9775. doi: 10.1038/s41467-024-52985-0.
  PUBMED    39532847
REFERENCE   2  (bases 1 to 5787)
  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: "Nat 
            Commun."; date: "2024-11-12"; volume: "15(1):9775. doi"; pages: " 
            10.1038/s41467-024-52985-0"
COMMENT     Sequence Label: OMM-long-CFAST10
FEATURES             Location/Qualifiers
     source          1..5787
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(1340..1358)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1341..1358)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1378..1395)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1384..1608
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1654..2082
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1741..1760)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1951..1972
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2091..2111)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2096..2425
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2276..2411
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2338..2357
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2492..3286
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2546..2565)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3156..3175
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3460..3581
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3497..3516)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3551..3570
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3630..3646)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3630..3646)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3643..3665)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3654..3670
                     /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)."
     promoter        complement(join(3678..3684,3685..3702,3703..3708))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3723..3744
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3861..3878)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4032..4620)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4112..4131)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4791..5582,5583..5651))
                     /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     5414..5433
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5652..5756)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttcgcaga
      901 ggacacttgt catggccgcc tctaaacctg tggaggcagc ggtggtcgca gccgctgtac
      961 cgagctccgg gagtggggtg ggcggcggcg ggactgcggg cccgggcacg ggggggctgc
     1021 cgcgatggca gctggctctg gcggtcgggg cacccctgct gctgggcgcg ggtgccatat
     1081 acctgtggag tcggcagcaa cggcgccggg aggccagagg ccggggcgac gccagcggcc
     1141 tgaagcgcaa cagcgaacgg aagaccccgg agggcagggc cagtccggcc ccgggcagcg
     1201 gacaccctga aggtcccggt gctcacttgg acatgaactc tcttggatcc ggtgacagct
     1261 attgggtctt tgtgaaacgg gaattctaag atatccatca cactggcggc cgctcgagca
     1321 tgcatctaga gggccctatt ctatagtgtc acctaaatgc tagagctcgc tgatcagcct
     1381 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     1441 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     1501 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     1561 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg
     1621 aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg
     1681 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     1741 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     1801 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     1861 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     1921 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     1981 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     2041 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg
     2101 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     2161 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     2221 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     2281 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     2341 ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt
     2401 ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc
     2461 tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
     2521 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     2581 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     2641 gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct
     2701 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     2761 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     2821 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     2881 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     2941 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
     3001 gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     3061 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     3121 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     3181 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     3241 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     3301 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc
     3361 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     3421 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     3481 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     3541 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     3601 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     3661 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     3721 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     3781 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     3841 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     3901 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     3961 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     4021 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     4081 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     4141 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     4201 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     4261 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4321 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4381 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4441 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     4501 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4561 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     4621 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     4681 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     4741 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     4801 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     4861 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     4921 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     4981 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     5041 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     5101 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     5161 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     5221 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     5281 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     5341 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     5401 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     5461 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     5521 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     5581 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     5641 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     5701 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     5761 tttccccgaa aagtgccacc tgacgtc
//
