LOCUS       sequence-226687-        8482 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent fusion protein used to visualise mitochondrial 
            nucleoids, with hygromycin selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8482)
  AUTHORS   McArthur K, Whitehead LW, Heddleston JM, Li L, Padman BS, Oorschot 
            V, Geoghegan ND, Chappaz S, Davidson S, San Chin H, Lane RM, 
            Dramicanin M, Saunders TL, Sugiana C, Lessene R, Osellame LD, Chew 
            TL, Dewson G, Lazarou M, Ramm G, Lessene G, Ryan MT, Rogers KL, van 
            Delft MF, Kile BT
  TITLE     BAK/BAX macropores facilitate mitochondrial herniation and mtDNA 
            efflux during apoptosis.
  JOURNAL   Science. 2018 Feb 23;359(6378). pii: 359/6378/eaao6047. doi: 
            10.1126/science.aao6047.
  PUBMED    29472455
REFERENCE   2  (bases 1 to 8482)
  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: "Science."; 
            date: "2018-02-23"; volume: "359(6378). pii: 359/6378/eaao6047. 
            doi"; pages: " 10.1126/science.aao6047"
COMMENT     Sequence Label: pMIH-TFAM-GFP
FEATURES             Location/Qualifiers
     source          1..8482
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    579..920
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             987..1403
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1333..1355
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1371..1387
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1416..1425
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2184..2897
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITLGMDELYK"
     CDS             2184..2894
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITLGMDELYK"
     primer_bind     complement(2223..2244)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2484..2503)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2831..2852
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    join(2918..3492,3493..3495,3496..3504)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3085..3102)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3312..3331
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     LTR             4883..5398
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     primer_bind     complement(5567..5583)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5567..5583)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5580..5602)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5591..5607
                     /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(5615..5621,5622..5639,5640..5645))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5660..5681
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5798..5815)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5969..6557)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6049..6068)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6728..7519,7520..7588))
                     /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     7351..7370
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7589..7693)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7761..7779
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7817..7839)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7939..7958
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8152..8174
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8302..8321
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(8482,1..514)
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat
       61 ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca
      121 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
      181 acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt
      241 ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc
      301 gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc
      361 ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt attcccaata
      421 aagcctcttg ctgtttgcat ccgaatcgtg gactcgctga tccttgggag ggtctcctca
      481 gattgattga ctgcccacct cgggggtctt tcatttggag gttccaccga gatttggaga
      541 cccctgccca gggaccaccg acccccccgc cgggaggtaa gctggccagc ggtcgtttcg
      601 tgtctgtctc tgtctttgtg cgtgtttgtg ccggcatcta atgtttgcgc ctgcgtctgt
      661 actagttagc taactagctc tgtatctggc ggacccgtgg tggaactgac gagttctgaa
      721 cacccggccg caaccctggg agacgtccca gggactttgg gggccgtttt tgtggcccga
      781 cctgaggaag ggagtcgatg tggaatccga ccccgtcagg atatgtggtt ctggtaggag
      841 acgagaacct aaaacagttc ccgcctccgt ctgaattttt gctttcggtt tggaaccgaa
      901 gccgcgcgtc ttgtctgctg cagcgctgca gcatcgttct gtgttgtctc tgtctgactg
      961 tgtttctgta tttgtctgaa aattagggcc agactgttac cactccctta agtttgacct
     1021 taggtcactg gaaagatgtc gagcggatcg ctcacaacca gtcggtagat gtcaagaaga
     1081 gacgttgggt taccttctgc tctgcagaat ggccaacctt taacgtcgga tggccgcgag
     1141 acggcacctt taaccgagac ctcatcaccc aggttaagat caaggtcttt tcacctggcc
     1201 cgcatggaca cccagaccag gtcccctaca tcgtgacctg ggaagccttg gcttttgacc
     1261 cccctccctg ggtcaagccc tttgtacacc ctaagcctcc gcctcctctt cctccatccg
     1321 ccccgtctct cccccttgaa cctcctcgtt cgaccccgcc tcgatcctcc ctttatccag
     1381 ccctcactcc ttctctaggc gccggaatta gatctgccac catggcgctg ttccggggaa
     1441 tgtggagcgt gctaaaagca ctggggcgca cgggggtcga gatgtgcgcg ggctgcgggg
     1501 gtcgcatccc ctcgtctatc agtcttgtct gtattccgaa gtgtttttcc agcatgggta
     1561 gctatccaaa gaaacctatg agttcatacc ttcgattttc cacagaacag ctacccaaat
     1621 ttaaagctaa acacccagat gcaaaacttt cagaattggt taggaaaatt gcagccctgt
     1681 ggagggagct accagaagca gaaaaaaagg tttatgaagc tgattttaaa gctgagtgga
     1741 aagcatacaa agaagctgtg agcaagtata aagagcagct aactccaagt cagctgatgg
     1801 gtatggagaa ggaggcccgg cagagacggt taaaaaagaa agcactggta aagagaagag
     1861 aattaatttt gcttggaaaa ccaaaaagac ctcgttcagc atataacatt tatgtatctg
     1921 aaagcttcca ggaggcaaag gatgattcgg ctcagggaaa attgaagctt gtaaatgagg
     1981 cttggaaaaa tctgtctcct gaggaaaagc aggcatatat tcagcttgct aaagatgata
     2041 ggattcgtta cgacaatgaa atgaagtctt gggaagagca gatggctgaa gttggacgaa
     2101 gtgatctcat ccgtcgaagt gtgaaacgat ccggagacat ctctgagcat gtcgacggta
     2161 ccgcgggccc gggatccacc ggtagcaagg gcgaggagct gttcaccggg gtggtgccca
     2221 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg
     2281 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc
     2341 ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct
     2401 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc
     2461 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt
     2521 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg
     2581 gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg
     2641 ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg
     2701 gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc
     2761 tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga
     2821 agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg
     2881 acgagctgta caagtaactc gaggttaacg aattccgccc ctctccctcc ccccccccta
     2941 acgttactgg ccgaagccgc ttggaataag gccggtgtgc gtttgtctat atgttatttt
     3001 ccaccatatt gccgtctttt ggcaatgtga gggcccggaa acctggccct gttttcttga
     3061 cgagcattcc taggggtctt tcccctctcg ccaaaggaat gcaaggtctg ttgaatgtcg
     3121 tgaaggaagc agttcctctg gaagcttctt gaagacaaac aacgtctgta gcgacccttt
     3181 gcaggcagcg gaacccccca cctggcgaca ggtgcctctg cggccaaaag ccacgtgtat
     3241 aagatacacc tgcaaaggcg gcacaacccc agtgccacgt tgtgagttgg atagttgtgg
     3301 aaagagtcaa atggctctcc tcaagcgtat tcaacaaggg gctgaaggat gcccagaagg
     3361 taccccattg tatgggatct gatctggggc ctcggtgcac atgctttaca tgtgtttagt
     3421 cgaggttaaa aaaacgtcta ggccccccga accacgggga cgtggttttc ctttgaaaaa
     3481 cacgatgata agcttgccac aacccgtacc aaagatggat agatccggaa agcctgaact
     3541 caccgcgacg tctgtcgaga agtttctgat cgaaaagttc gacagcgtct ccgacctgat
     3601 gcagctctcg gagggcgaag aatctcgtgc tttcagcttc gatgtaggag ggcgtggata
     3661 tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa gatcgttatg tttatcggca
     3721 ctttgcatcg gccgcgctcc cgattccgga agtgcttgac attggggaat tcagcgagag
     3781 cctgacctat tgcatctccc gccgtgcaca gggtgtcacg ttgcaagacc tgcctgaaac
     3841 cgaactgccc gctgttctgc agccggtcgc ggaggccatg gatgcgatcg ctgcggccga
     3901 tcttagccag acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc aatacactac
     3961 atggcgtgat ttcatatgcg cgattgctga tccccatgtg tatcactggc aaactgtgat
     4021 ggacgacacc gtcagtgcgt ccgtcgcgca gctctcgatg agctgatgct ttgggccgag
     4081 gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg
     4141 gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa
     4201 tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg
     4261 cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg
     4321 ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca
     4381 gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt
     4441 acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc
     4501 gatagtggaa accgacgccc cagcactcgt ccgagggcaa aggaatagag tagatgccga
     4561 ccgaacaaga gctgatttcg agaacgcctc agccagcaac tcgcgcgagc ctagcaaggc
     4621 aaatgcgaga gaacggcctt acgcttggtg gcacagttct cgtccacagt tcgctaagct
     4681 cgctcggctg ggtcgcggga gggccggtcg cagtgattca ggcccttctg gattgtgttg
     4741 gtccccaggg cacgattgtc atgcccacgc actcgggtga tctgactgat cccgcagatt
     4801 ggagatcgcc gcccgtgcct gccgattggg tgcagatccg ataaaataaa agattttatt
     4861 tagtctccag aaaaaggggg gaatgaaaga ccccacctgt aggtttggca agctagctta
     4921 agtaacgcca ttttgcaagg catggaaaat acataactga gaatagagaa gttcagatca
     4981 aggttaggaa cagagagaca gcagaatatg ggccaaacag gatatctgtg gtaagcagtt
     5041 cctgccccgg ctcagggcca agaacagatg gtccccagat gcggtcccgc cctcagcagt
     5101 ttctagagaa ccatcagatg tttccagggt gccccaagga cctgaaaatg accctgtgcc
     5161 ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc tgctccccga
     5221 gctcaataaa agagcccaca acccctcact cggcgcgcca gtcctccgat agactgcgtc
     5281 gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt gtggtctcgc
     5341 tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg tctttcatgg
     5401 gtaacagttt cttgaagttg gagaacaaca ttctgagggt aggagtcgaa tattaagtaa
     5461 tcctgactca attagccact gttttgaatc cacatactcc aatactcctg aaatagttca
     5521 ttatggacag cgcagaagag ctggggagaa ttaattcgta atcatggtca tagctgtttc
     5581 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     5641 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     5701 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     5761 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     5821 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     5881 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     5941 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     6001 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     6061 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     6121 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     6181 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     6241 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     6301 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     6361 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     6421 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     6481 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     6541 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     6601 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     6661 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     6721 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     6781 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     6841 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     6901 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     6961 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     7021 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     7081 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     7141 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     7201 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     7261 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     7321 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     7381 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     7441 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     7501 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     7561 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     7621 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     7681 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta
     7741 tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg
     7801 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     7861 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     7921 ggggctggct taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt
     7981 gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgccat tcgccattca
     8041 ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg
     8101 cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac
     8161 gacgttgtaa aacgacggcg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc
     8221 cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg
     8281 gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt
     8341 ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg cgattagtcc aatttgttaa
     8401 agacaggata tcagtggtcc aggctctagt tttgactcaa caatatcacc agctgaagcc
     8461 tatagagtac gagccataga ta
//
