LOCUS       sequence-379624-        8493 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To produce mutants mimicking phosphorylated Mfn1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8493)
  AUTHORS   Pyakurel A, Savoia C, Hess D, Scorrano L
  TITLE     Extracellular regulated kinase phosphorylates mitofusin 1 to control
            mitochondrial morphology and apoptosis.
  JOURNAL   Mol Cell. 2015 Apr 16;58(2):244-54. doi: 
            10.1016/j.molcel.2015.02.021. Epub 2015 Mar 19.
  PUBMED    25801171
REFERENCE   2  (bases 1 to 8493)
  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"; doi: 
            "10.1016/j.molcel.2015.02.021"; journalName: "Mol Cell"; date: 
            "2015-04-16- 16"; volume: "58"; issue: "2"; pages: "244-54"
COMMENT     Sequence Label: p3xFLAG-CMV-14-Mfn1-T562D
FEATURES             Location/Qualifiers
     source          1..8493
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    160..294
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(197..216)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     251..270
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(299..318)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(300..318)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(332..348)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(332..348)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(345..367)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    356..372
                     /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(380..386,387..404,405..410))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    425..446
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(563..580)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(734..1322)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(814..833)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1493..2284,2285..2353))
                     /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     2116..2135
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2354..2458)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2485..2940
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2572..2591)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2782..2803
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3066..3088
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3080..3097
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3081..3097
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        3258..3637
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3638..3841
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3792..3812
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             6116..6181
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    6192..6814
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(6391..6410)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(6838..6858)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6856..7153
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             7207..8001
                     /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(7261..7280)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7871..7890
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 aagctggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gccgatcata
       61 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
      121 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
      181 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
      241 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg gatcaattcc
      301 ctatagtgag tcgtattaaa ttcgtaatca tgtcatagct gtttcctgtg tgaaattgtt
      361 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
      421 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
      481 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
      541 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
      601 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
      661 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
      721 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
      781 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
      841 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
      901 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
      961 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     1021 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     1081 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     1141 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     1201 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     1261 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     1321 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     1381 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     1441 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     1501 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     1561 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     1621 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     1681 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     1741 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     1801 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     1861 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     1921 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     1981 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     2041 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     2101 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     2161 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     2221 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     2281 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     2341 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     2401 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     2461 catttccccg aaaagtgcca cctgacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     2521 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     2581 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     2641 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     2701 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     2761 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     2821 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     2881 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt
     2941 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     3001 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
     3061 gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagctgatct atacattgaa
     3121 tcaatattgg caattagcca tattagtcat tggttatata gcataaatca atattggcta
     3181 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
     3241 aatatgaccg ccatgttgac attgattatt gactagttat taatagtaat caattacggg
     3301 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
     3361 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
     3421 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
     3481 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
     3541 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
     3601 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
     3661 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
     3721 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
     3781 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
     3841 tcgtttagtg aaccgtcaga attaagcttc caccatggca gaaacggtat ctccactgaa
     3901 gcacttcgtg ctggcaaaga aggccatcac tgcgatcttc ggccagttac tggagtttgt
     3961 tactgagggc tcacattttg ttgaagcaac atacaggaac ccggaacttg atcggatagc
     4021 aaccgaggat gatctggtgg agatacaggg ctacagaaac aagctcgctg tcattgggga
     4081 ggtgctgtct cggaggcata tgaaagtggc attttttggc aggacaagta gtggcaagag
     4141 ctctgtcatc aacgcaatgc tgtgggataa agtcctcccc agcgggattg gtcacacaac
     4201 caactgcttc ctgagtgtgg agggaactga tggagataaa gcctacctta tgaccgaagg
     4261 gtcagatgag aagaaaagcg tgaagactgt taatcagctg gcccatgccc tccatatgga
     4321 taaagacttg aaagctggtt gtcttgtgca cgtattttgg cccaaagcaa aatgtgccct
     4381 cttgagagat gacctggttt tagtagacag cccaggtaca gatgtcacca cagagctgga
     4441 catctggatt gataagttct gccttgatgc tgatgttttt gttttggtgg caaactcgga
     4501 atcaacgctg atgaacacgg agaaacattt tttccataag gtgaatgagc ggctctccaa
     4561 gccaaacatt tttatcctga ataatcgttg ggatgcttct gcttcagagc cagaatatat
     4621 ggaggatgtg cgcagacagc acatggaaag atgtcttcac ttcttggtag aagagctcaa
     4681 ggttgtgagt cctttagaag ctcggaatcg gatctttttt gtttcagcca aggaagttct
     4741 caactccaga atgaataaag cccaggggat gccagaaggt ggaggggcac ttgcagaagg
     4801 atttcaggca agattacagg agtttcaaaa ttttgaacaa acttttgagg agtgtatctc
     4861 gcagtcagca gtgaaaacaa agtttgaaca gcacactatc agagctaaac agatactaga
     4921 cactgtgaaa aacatactgg actcagtaaa cgtggcagca gcagagaaga gggtttattc
     4981 aatggaagag agggaagacc aaatcgacag actggacttt attcgaaacc agatgaacct
     5041 tttaacaatg gatgttaaga agaagatcaa ggaggtcacg gaggaggtgg caaacaaggt
     5101 ttcttgtgca atgacagatg agatttgtcg cctgtctgtt ttggttgatg agttttgttc
     5161 tgagtttcat cctaccccca gtgtgctgaa agtgtataag agtgaattaa ataagcacat
     5221 agaagatggc atgggaagaa atttggctga tcggtgcacc agtgaagtca atgcctccat
     5281 tctccaatct cagcaagaaa tcattgaaaa tttgaagcca ctacttccag ctggtataca
     5341 gaataaactt catacattaa tcccttgcaa aaagtttgat ctcagctatg atctcaattg
     5401 ccacaagctg tgttcggatt ttcaagaaga cattgtgttt cggttttccc tgggctggtc
     5461 ttcccttgta catcgattcc tgggttctac gaatgcacag agggtgctgc ttgggctgtc
     5521 agagcctatc tttcaggtcc ctagatcttt agcttcagac cctactgctc cttctaaccc
     5581 agcagcacca gataatgcag cccaggaaga gctcatgatc acactgatca caggattggc
     5641 atccctcacg tctagaacct ccatgggcat catagttgtt gggggcgtga tttggaaaac
     5701 agtgggctgg aaactcatct ctgtcacctt aagtatgtat ggagctttgt acctttatga
     5761 aagactgact tggactactc gtgcgaaaga gagagcgttt aagcagcagt ttgtaaacta
     5821 tgcaactgaa aaactgcaga tgattgtgaa gttcacaagt gcaaactgca gccaccaagt
     5881 ccagcaagaa atggccacca cttttgctcg actgtgccaa caagttgatg ttactcagaa
     5941 acatctggaa gaagaaattg ctagattatc caaagagatt gaccaactgg agaaaataca
     6001 aaacaattca aagctcttaa gaaataaagc tgttcaactt gaacgtgagc tggaaaattt
     6061 ttcgaagcag tttctccacc cgagcagtgg agaatcccct agaggatccc gggctgacta
     6121 caaagaccat gacggtgatt ataaagatca tgacatcgac tacaaggatg acgatgacaa
     6181 gtagtgatcc cgggtggcat ccctgtgacc cctccccagt gcctctcctg gccctggaag
     6241 ttgccactcc agtgcccacc agccttgtcc taataaaatt aagttgcatc attttgtctg
     6301 actaggtgtc cttctataat attatggggt ggaggggggt ggtatggagc aaggggcaag
     6361 ttgggaagac aacctgtagg gcctgcgggg tctattggga accaagctgg agtgcagtgg
     6421 cacaatcttg gctcactgca atctccgcct cctgggttca agcgattctc ctgcctcagc
     6481 ctcccgagtt gttgggattc caggcatgca tgaccaggct cagctaattt ttgttttttt
     6541 ggtagagacg gggtttcacc atattggcca ggctggtctc caactcctaa tctcaggtga
     6601 tctacccacc ttggcctccc aaattgctgg gattacaggc gtgaaccact gctcccttcc
     6661 ctgtccttct gattttaaaa taactatacc agcaggagga cgtccagaca cagcataggc
     6721 tacctggcca tgcccaaccg gtgggacatt tgagttgctt gcttggcact gtcctctcat
     6781 gcgttgggtc cactcagtag atgcctgttg aattgggtac gcggccagct tggctgtgga
     6841 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     6901 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     6961 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     7021 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     7081 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     7141 gaggcttttt tggaggaatt gatcagcttg ggatctgatc aagagacagg atgaggatcg
     7201 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
     7261 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
     7321 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
     7381 gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
     7441 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
     7501 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
     7561 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
     7621 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
     7681 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg
     7741 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
     7801 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
     7861 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
     7921 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
     7981 cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc
     8041 ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     8101 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     8161 tcttcgccca ccccgggctc gatcccctcg cgagttggtt cagctgctgc ctgaggctgg
     8221 acgacctcgc ggagttctac cggcagtgca aatccgtcgg catccaggaa accagcagcg
     8281 gctatccgcg catccatgcc cccgaactgc aggagtgggg aggcacgatg gccgctttgg
     8341 tcgacccgga cgggacgctc ctgcgcctga tacagaacga attgcttgca ggcatctcat
     8401 gagtgtgtct tcccgttttc cgcctgaggt cactgcgtgg atggagcgct ggcgcctgct
     8461 gcgcgacggc gagctgctca ccacccactc gcc
//
