LOCUS       sequence-484682-        5615 bp    DNA     circular SYN 26-AUG-2025
DEFINITION  endogenously tags MFN2 with Halo tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5615)
  AUTHORS   Thayer JA, Petersen JD, Huang X, Hawrot J, Ramos DM, Sekine S, Li Y,
            Ward ME, Narendra DP
  TITLE     Novel reporter of the PINK1-Parkin mitophagy pathway identifies its 
            damage sensor in the import gate.
  JOURNAL   bioRxiv [Preprint]. 2025 Feb 20:2025.02.19.639160. doi: 
            10.1101/2025.02.19.639160.
  PUBMED    40027798
REFERENCE   2  (bases 1 to 5615)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 26, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2025 Feb 20:2025.02.19.639160. doi: 
            10.1101/2025.02.19.639160."
COMMENT     Sequence Label: MFN2-Halo donor plasmid
FEATURES             Location/Qualifiers
     source          1..5615
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             761..1648
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="AEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSY
                     VWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIH
                     DWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIID
                     QNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVE
                     EYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLI
                     GSEIARWLSTLEISG"
     primer_bind     complement(2699..2715)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2699..2715)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2712..2734)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2723..2739
                     /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(2747..2753,2754..2771,2772..2777))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2792..2813
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2930..2947)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3101..3689)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3181..3200)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3860..4651,4652..4720))
                     /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     4483..4502
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4721..4825)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4893..4911
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     5071..5090
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5284..5306
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5298..5315
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5299..5315
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 agcacacggg gaccctgcag aagtcctcgg cctgcctagc ccatgatagc atcagcacac
       61 ggggaccctg cacttcctgc cctcgcccct gacgtcagag tctgcagggt ggcagagcca
      121 gaggggagat ggatcagagg gtgtcagctc caaccctcac cggatcactg ggctttccct
      181 gccctcgact gcggatgtgc cccccccgca cccccacctg gggacagctg cttcctaggg
      241 acacactgct tgccctctgg ggactctcag aaagggggaa tggggggaac tgctgcttgt
      301 cagccacagt gacacagctg aacctggaaa gggtctcctg tcagcacctc aaccccttgc
      361 ttggcaagca gggaaactga gttcccgaca gctcagggac ttgcacatgg tcctgggtga
      421 aggaaacaca gggcttgaga atcagaaaag cctaacttgt agctccctgg cttattagcc
      481 gtgtggcctt gggaagttat ttaacccctc tgggcctcct tttccccatc tgtgagacag
      541 ggataaacac gattgttgga ggatgatgta agggtgtgtg tcaagcgtcc ttaggatggt
      601 gcctggcggg tagtcctaat actgcctatc atcagctatc atggttacaa aagaaccatt
      661 tctttgcagg aataaagccg gttggttgga cagtgagctc aacatgttca cacatcagta
      721 tctgcagcca tcccgtggag gatcgggagg atcaggatcc gcagaaatcg gtactggctt
      781 tccattcgac ccccattatg tggaagtcct gggcgagcgc atgcactacg tcgatgttgg
      841 tccgcgcgat ggcacccctg tgctgttcct gcacggtaac ccgacctcct cctacgtgtg
      901 gcgcaacatc atcccgcatg ttgcaccgac ccatcgctgc attgctccag acctgatcgg
      961 tatgggcaaa tccgacaaac cagacctggg ttatttcttc gacgaccacg tccgcttcat
     1021 ggatgccttc atcgaagccc tgggtctgga agaggtcgtc ctggtcattc acgactgggg
     1081 ctccgctctg ggtttccact gggccaagcg caatccagag cgcgtcaaag gtattgcatt
     1141 tatggagttc atccgcccta tcccgacctg ggacgaatgg ccagaatttg cccgcgagac
     1201 cttccaggcc ttccgcacca ccgacgtcgg ccgcaagctg atcatcgatc agaacgtttt
     1261 tatcgagggt acgctgccga tgggtgtcgt ccgcccgctg actgaagtcg agatggacca
     1321 ttaccgcgag ccgttcctga atcctgttga ccgcgagcca ctgtggcgct tcccaaacga
     1381 gctgccaatc gccggtgagc cagcgaacat cgtcgcgctg gtcgaagaat acatggactg
     1441 gctgcaccag tcccctgtcc cgaagctgct gttctggggc accccaggcg ttctgatccc
     1501 accggccgaa gccgctcgcc tggccaaaag cctgcctaac tgcaaggctg tggacatcgg
     1561 cccgggtctg aatctgctgc aagaagacaa cccggacctg atcggcagcg agatcgcgcg
     1621 ctggctgtcg acgctcgaga tttccggcta ggatatctgg gcacctgagg cggagtctgc
     1681 gtggagaggg gcggtgctgc cagccctaag tgccatgtgg gctcccccag gggcacgtgt
     1741 ggctcctgcc ccctggccac tgccaagaga atgaagcacc cagtctcgta ccattttgag
     1801 ccctccagca ctacttattt tcccccacct ttgcctgctg ttgctggaag agctggctca
     1861 tacccccaaa ggacactttc agcgacagct atggacagca tggtaccaag gagttaagtt
     1921 gaggcttttt ccagctttct ctggttcatt tgattgcttg ataaggcctc aggatctcag
     1981 cattgcacaa tgcctcatgg aagcctttga gggtatcaca cagacacccc caccttcctc
     2041 cagcctgtgc gcacctgccc tccttgcagc ccagcacacc tgcaggtgta agggacgatt
     2101 ggagtttctt cccagagagt ctgtcccaga aggactgtgg cttgtgtgtg tccatctcgc
     2161 ctgttggctc agtgcttcat cccatttgca gagcctcaga cacgtcttgg tggtgaggct
     2221 cagttacccc tgggcttagg ctgaggcggg ccctgtgctg ggggtggtag aaaggatgct
     2281 gctgaggcag ctggaggagt gggagtagct cagaggggag ggctgttgga tgtatgggga
     2341 gctggcagag caggtggcag tcactgggac aaggagggac ttgcctctct tctcattatt
     2401 gtgtcctttg ctttagtgtc agtcctggac ttgtgcaggc ctgttttgtg tagatctgtt
     2461 ttggaagatg gcatggtcta ggtggttgaa ggatgtagta gaaggatgga tggtggaagg
     2521 tggggacgtt ggtggctggc tgaggtgcat gggccccaca caggacagct ggagaatggg
     2581 ccgtccactt ggcctcgttc tgcgaggggc tcatgggtct gagagccccc acccactagg
     2641 cttgattgca tccctgtgtc gactgcagag gcctgcatgc aagcttggcg taatcatggt
     2701 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     2761 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     2821 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     2881 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     2941 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     3001 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     3061 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     3121 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     3181 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     3241 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     3301 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     3361 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     3421 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     3481 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     3541 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     3601 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     3661 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     3721 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     3781 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     3841 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     3901 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     3961 agggcttacc atctggcccc agtgctgcaa tgataccgcg agatccacgc tcaccggctc
     4021 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     4081 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     4141 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     4201 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     4261 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     4321 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     4381 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     4441 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     4501 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     4561 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     4621 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     4681 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     4741 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     4801 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag
     4861 aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc
     4921 ttgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg tagacggtca
     4981 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
     5041 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
     5101 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
     5161 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
     5221 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
     5281 ttacccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcggt acctcgcgaa
     5341 tgcatctaga tactggcaga gctgggattg gaactcagat ctccatgggg tctctgtgga
     5401 gctttcatta ataatttgct ggaaagaaca tcagtctgag gagtctggag tcctgggttc
     5461 tctccagcca ctcatgtttg gcccaagctc ttaactctcc tgtgtctcat tattttcatc
     5521 tgtaaaatgg gagtaagatt tcatgacaca gctgcatgac tgaatgtaaa agtgccctgc
     5581 aaaagtcctc ggcctgccca gctcatgata acatc
//
