LOCUS       sequence-393506-        6555 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transiently expresses mitochondria localized TurboID with V5 tags.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6555)
  AUTHORS   Yan T, Julio AR, Villanueva M, Jones AE, Ball AB, Boatner LM, Turmon
            AC, Nguyen KB, Yen SL, Desai HS, Divakaruni AS, Backus KM
  TITLE     Proximity-labeling chemoproteomics defines the subcellular 
            cysteinome and inflammation-responsive mitochondrial redoxome.
  JOURNAL   Cell Chem Biol. 2023 Jul 20;30(7):811-827.e7. doi: 
            10.1016/j.chembiol.2023.06.008. Epub 2023 Jul 6.
  PUBMED    37419112
REFERENCE   2  (bases 1 to 6555)
  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.chembiol.2023.06.008"; journalName: "Cell Chem Biol"; 
            date: "2023-07-20- 20"; volume: "30"; issue: "7"; pages: 
            "811-827.e7"
COMMENT     Sequence Label: Mito-TurboID
FEATURES             Location/Qualifiers
     source          1..6555
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        115..494
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        495..698
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     649..669
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     743..762
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        743..761
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             936..1892
                     /codon_start=1
                     /product="mutant of the E. coli biotin ligase BioID 
                     engineered for improved catalytic efficiency (Branon et 
                     al., 2018)"
                     /label=TurboID
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIPLLNAKQILGQLDGGSVAVLPVVDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGSRGRKWFSPFGANLYLSMFWRLKRGPAAIGLGPVIGIVMAEALRKLG
                     ADKVRVKWPNDLYLQDRKLAGILVELAGITGDAAQIVIGAGINVAMRRVEESVVNQGWI
                     TLQEAGINLDRNTLAATLIRELRAALELFEQEGLAPYLPRWEKLDNFINRPVKLIIGDK
                     EIFGISRGIDKQGALLLEQDGVIKPWMGGEISLRSAEK"
     CDS             1899..1940
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     promoter        complement(1988..2006)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1989..2006)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2026..2043)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2032..2256
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2302..2730
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2389..2408)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2599..2620
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2739..2759)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2744..3073
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2924..3059
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2986..3005
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3140..3934
                     /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(3194..3213)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3804..3823
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4108..4229
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4145..4164)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4199..4218
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4278..4294)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4278..4294)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4291..4313)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4302..4318
                     /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(4326..4332,4333..4350,4351..4356))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4371..4392
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4509..4526)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4680..5268)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4760..4779)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5439..6230,6231..6299))
                     /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"
     promoter        complement(6300..6404)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6479..6498)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
       61 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      121 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      181 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      241 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      301 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      361 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      421 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      481 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      541 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      601 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      661 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      721 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgggccg
      781 ccaccatgct ggccacccgc gtgttcagcc tggtgggcaa gcgcgccatc agcaccagcg
      841 tgtgcgtgcg cgcccacgga tcaggaagcg gcggccgcgg atcaggaagc ggatcaggaa
      901 gcggatcagg aagcggatca ggaagcggat caggaaaaga caatactgtg cctctgaagc
      961 tgatcgctct cctggctaat ggcgagttcc atagtggcga acagctggga gaaaccctgg
     1021 gcatgtccag ggccgctatc aacaagcaca ttcagactct gcgcgactgg ggcgtggacg
     1081 tgttcaccgt gcccggaaag ggctactctc tgcccgagcc tatcccgctg ctgaacgcta
     1141 aacagattct gggacagctg gacggcggga gcgtggcagt cctgcctgtg gtcgactcca
     1201 ccaatcagta cctgctggat cgaatcggcg agctgaagag tggggatgct tgcattgcag
     1261 aatatcagca ggcagggaga ggaagcagag ggaggaaatg gttctctcct tttggagcta
     1321 acctgtacct gagtatgttt tggcgcctga agcggggacc agcagcaatc ggcctgggcc
     1381 cggtcatcgg aattgtcatg gcagaagcgc tgcgaaagct gggagcagac aaggtgcgag
     1441 tcaaatggcc caatgacctg tatctgcagg atagaaagct ggcaggcatc ctggtggagc
     1501 tggccggaat aacaggcgat gctgcacaga tcgtcattgg cgccgggatt aacgtggcta
     1561 tgaggcgcgt ggaggaaagc gtggtcaatc agggctggat cacactgcag gaagcaggga
     1621 ttaacctgga caggaatact ctggccgcta cgctgatccg agagctgcgg gcagccctgg
     1681 aactgttcga gcaggaaggc ctggctccat atctgccacg gtgggagaag ctggataact
     1741 tcatcaatag acccgtgaag ctgatcattg gggacaaaga gattttcggg attagccggg
     1801 ggattgataa acagggagcc ctgctgctgg aacaggacgg agttatcaaa ccctggatgg
     1861 gcggagaaat cagtctgcgg tctgccgaaa aggctagcgg caagcccatc cccaaccccc
     1921 tgctgggcct ggacagcacc taatgaggat ccggcaagcc catccccaac cccctgctgg
     1981 gccctattct atagtgtcac ctaaatgcta gagctcgctg atcagcctcg actgtgcctt
     2041 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
     2101 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
     2161 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca
     2221 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct
     2281 ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     2341 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     2401 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     2461 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     2521 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     2581 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     2641 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     2701 agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg
     2761 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2821 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     2881 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     2941 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     3001 cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     3061 aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcaagagac
     3121 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
     3181 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
     3241 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
     3301 cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg
     3361 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
     3421 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
     3481 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
     3541 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
     3601 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
     3661 caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
     3721 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
     3781 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     3841 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     3901 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc
     3961 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
     4021 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     4081 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa
     4141 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     4201 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag
     4261 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     4321 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     4381 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     4441 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     4501 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     4561 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     4621 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     4681 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     4741 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     4801 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     4861 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     4921 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     4981 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     5041 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     5101 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     5161 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     5221 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     5281 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     5341 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     5401 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     5461 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     5521 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     5581 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     5641 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     5701 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     5761 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     5821 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     5881 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     5941 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     6001 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     6061 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     6121 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     6181 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     6241 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     6301 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     6361 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     6421 gtgccacctg acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta
     6481 caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg
     6541 tcgctgagta gtgcg
//
