LOCUS       sequence-361505-        8684 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mitochondrial labile heme reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8684)
  AUTHORS   Martinez-Guzman O, Willoughby MM, Saini A, Dietz JV, Bohovych I, 
            Medlock AE, Khalimonchuk O, Reddi AR
  TITLE     Mitochondrial-nuclear heme trafficking in budding yeast is regulated
            by GTPases that control mitochondrial dynamics and ER contact sites.
  JOURNAL   J Cell Sci. 2020 May 20;133(10). pii: jcs.237917. doi: 
            10.1242/jcs.237917.
  PUBMED    32265272
REFERENCE   2  (bases 1 to 8684)
  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: "J Cell 
            Sci."; date: "2020-05-20"; volume: "133(10). pii: jcs.237917. doi"; 
            pages: " 10.1242/jcs.237917"
COMMENT     Sequence Label: mito-yHS1-M7A,H102A
FEATURES             Location/Qualifiers
     source          1..8684
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        432..536
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(537..605,606..1397)
                     /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     complement(755..774)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1568..2156
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2057..2076
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2310..2327
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2444..2465
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2480..2485,2486..2503,2504..2510)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2518..2534
                     /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)."
     primer_bind     2523..2545
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2542..2558
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2542..2558
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2577..2597
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2579..2597
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2622..3265
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     3159..3182
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             3283..3348
                     /codon_start=1
                     /product="mitochondrial presequence of yeast cytochrome c 
                     oxidase subunit IV (Hurt et al., 1984)"
                     /label=pre-CoxIV
                     /translation="MLSLRQSIRFFKPATRTLCSSR"
     terminator      5134..5381
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(5150..5168)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(5399..5418)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5400..5418)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5428..5445)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5428..5444)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5437..5459)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5585..6040
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5672..5691)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5882..5903
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6341..6748
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             6749..7843
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     primer_bind     complement(8337..8356)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8456..8478
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(8516..8534)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    join(8576..8684,1..395)
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
ORIGIN
        1 gtaaataaag aaggtagaag agttacggaa tgaagaaaaa aaaataaaca aaggtttaaa
       61 aaatttcaac aaaaagcgta ctttacatat atatttatta gacaagaaaa gcagattaaa
      121 tagatataca ttcgattaac gataagtaaa atgtaaaatc acaggatttt cgtgtgtggt
      181 cttctacaca gacaagatga aacaattcgg cattaatacc tgagagcagg aagagcaaga
      241 taaaaggtag tatttgttgg cgatccccct agagtctttt acatcttcgg aaaacaaaaa
      301 ctattttttc tttaatttct ttttttactt tctattttta atttatatat ttatattaaa
      361 aaatttaaat tataattatt tttatagcac gtgatgaaaa ggacccaggt ggcacttttc
      421 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
      481 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
      541 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
      601 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
      661 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
      721 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
      781 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
      841 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
      901 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
      961 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     1021 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     1081 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     1141 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     1201 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     1261 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     1321 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     1381 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     1441 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     1501 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     1561 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     1621 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     1681 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     1741 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     1801 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     1861 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     1921 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     1981 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     2041 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     2101 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     2161 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     2221 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     2281 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     2341 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     2401 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca
     2461 ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg
     2521 tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc aagcgcgcaa
     2581 ttaaccctca ctaaagggaa caaaagctgg agctcagttt atcattatca atactcgcca
     2641 tttcaaagaa tacgtaaata attaatagta gtgattttcc taactttatt tagtcaaaaa
     2701 attagccttt taattctgct gtaacccgta catgcccaaa atagggggcg ggttacacag
     2761 aatatataac atcgtaggtg tctgggtgaa cagtttattc ctggcatcca ctaaatataa
     2821 tggagcccgc tttttaagct ggcatccaga aaaaaaaaga atcccagcac caaaatattg
     2881 ttttcttcac caaccatcag ttcataggtc cattctctta gcgcaactac agagaacagg
     2941 ggcacaaaca ggcaaaaaac gggcacaacc tcaatggagt gatgcaacct gcctggagta
     3001 aatgatgaca caaggcaatt gacccacgca tgtatctatc tcattttctt acaccttcta
     3061 ttaccttctg ctctctctga tttggaaaaa gctgaaaaaa aaggttgaaa ccagttccct
     3121 gaaattattc ccctacttga ctaataagta tataaagacg gtaggtattg attgtaattc
     3181 tgtaaatcta tttcttaaac ttcttaaatt ctacttttat agttagtctt ttttttagtt
     3241 ttaaaacacc agaacttagt ttcgacggat tctagaacta gtatgctttc actacgtcaa
     3301 tctataagat ttttcaagcc agccacaaga actttgtgta gctctagata tctgcttaag
     3361 cttcatatgc acatggtatc ggaactgatc aaggaaaaca tgcacatgaa gctgtatatg
     3421 gaaggcacgg tcaacaacca ccactttaaa tgcacctcag aaggcgaagg taaaccttat
     3481 gaaggtaccc aaactatgag aattaaggct gtcgaaggtg gcccactgcc gtttgccttc
     3541 gatatcttag cgacttcttt tatgtacggc tcaaagacat tcatcaacca tacgcaaggt
     3601 atcccagatt tctttaaaca gtcctttccg gaaggcttca cctgggaacg cgttaccact
     3661 tatgaagacg gtggcgtgct gacagctacg caagatactt cgttacagga cggttgtttg
     3721 atttacaacg tcaaaatccg tggcgtaaat tttcctagca acggtccagt catgcagaaa
     3781 aagacattag gctgggaagc tagtaccgaa actttgtatc cggcagatgg tggcctggaa
     3841 ggtcgcgctg acatggcact gaaattagta ggtggcggtc acttgatttg caatctgaag
     3901 acaacgtacc gtagtaaaaa gccggccaaa aacttaaaga tgcctggtgt ttattacgtg
     3961 gatcgtagat tggaaagaat caaagaagcg gacaaggaaa cgtatgtgga acagcacgaa
     4021 gtcgccgtcg cacgctactg tgatttaccg tccaagttag gtcatagagg tggatccatg
     4081 gtgtccaaag gtgaagaact gtttactggt gtagtgccga ttttagttga attagacggt
     4141 gatgtgaatg gtcataagtt tagcgtgtcg ggtgaaggcg aaggtgatgc tacctttggt
     4201 ggctctgcag atctggaaga caacgcggaa actctgaacg ataacttgaa ggttatcgaa
     4261 aaggccgata atgctgcaca agtgaaagac gcgttaacaa agatgcgcgc cgcggctttg
     4321 gatgctcaga aagcaacgcc accgaaactg gaggataagt ctcctgactc accagaaatg
     4381 aaagattttc gtcatggctt cgacatttta gtgggtcaaa tcgatgacgc cttgaaactg
     4441 gcgaacgagg gtaaagtcaa ggaagctcaa gcagccgcgg aacagctgaa gaccactcgt
     4501 aatgcatatg ctcagaaata cagaggtggc aaactgacct tgaagtttat ctgtacaacg
     4561 ggtaaactgc ctgttccatg gccgactttg gtgaccactc tgggctatgg tgtacaatgc
     4621 ttctcacgtt acccagatca tatgaaacag cacgactttt tcaagagcgc catgccggaa
     4681 ggctacgttc aagaacgcac tattttcttt aaggatgacg gtaactacaa gacacgtgcg
     4741 gaagtcaaat ttgaaggcga tacgctggta aacagaatcg aattgaaggg tatcgatttc
     4801 aaggaagacg gcaacatctt gggtcataaa ttggaataca actacaactc ccacaacgtc
     4861 tacatcatgg ctgataagca aaagaatggc atcaaggtaa acttcaagat cagacataac
     4921 atcgaagatg gttctgttca gctggctgac cactatcaac agaatacacc tattggcgat
     4981 ggtccggtgc tgttacctga taaccattac ttgagcacgc agagtgcact gtccaaagat
     5041 ccaaatgaaa agcgcgacca catggtcttg ttggaatttg tcaccgctgc tggtatcact
     5101 ttgggtatgg atgaactgta taaatgactc gagtcatgta attagttatg tcacgcttac
     5161 attcacgccc tccccccaca tccgctctaa ccgaaaagga aggagttaga caacctgaag
     5221 tctaggtccc tatttatttt tttatagtta tgttagtatt aagaacgtta tttatatttc
     5281 aaatttttct tttttttctg tacagacgcg tgtacgcatg taacattata ctgaaaacct
     5341 tgcttgagaa ggttttggga cgctcgaagg ctttaatttg cggccggtac ccaattcgcc
     5401 ctatagtgag tcgtattacg cgcgctcact ggccgtcgtt ttacaacgtc gtgactggga
     5461 aaaccctggc gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg
     5521 taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga
     5581 atggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     5641 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     5701 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     5761 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     5821 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     5881 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     5941 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     6001 tttaacgcga attttaacaa aatattaacg cttacaattt cctgatgcgg tattttctcc
     6061 ttacgcatct gtgcggtatt tcacaccgca tatcgaccgg tcgaggagaa cttctagtat
     6121 atctacatac ctaatattat tgccttatta aaaatggaat cccaacaatt acatcaaaat
     6181 ccacattctc ttcaaaatca attgtcctgt acttccttgt tcatgtgtgt tcaaaaacgt
     6241 tatatttata ggataattat actctatttc tcaacaagta attggttgtt tggccgagcg
     6301 gtctaaggcg cctgattcaa gaaatatctt gaccgcagtt aactgtggga atactcaggt
     6361 atcgtaagat gcaagagttc gaatctctta gcaaccatta tttttttcct caacataacg
     6421 agaacacaca ggggcgctat cgcacagaat caaattcgat gactggaaat tttttgttaa
     6481 tttcagaggt cgcctgacgc atataccttt ttcaactgaa aaattgggag aaaaaggaaa
     6541 ggtgagagcg ccggaaccgg cttttcatat agaatagaga agcgttcatg actaaatgct
     6601 tgcatcacaa tacttgaagt tgacaatatt atttaaggac ctattgtttt ttccaatagg
     6661 tggttagcaa tcgtcttact ttctaacttt tcttaccttt tacatttcag caatatatat
     6721 atatatattt caaggatata ccattctaat gtctgcccct aagaagatcg tcgttttgcc
     6781 aggtgaccac gttggtcaag aaatcacagc cgaagccatt aaggttctta aagctatttc
     6841 tgatgttcgt tccaatgtca agttcgattt cgaaaatcat ttaattggtg gtgctgctat
     6901 cgatgctaca ggtgttccac ttccagatga ggcgctggaa gcctccaaga aggctgatgc
     6961 cgttttgtta ggtgctgtgg gtggtcctaa atggggtacc ggtagtgtta gacctgaaca
     7021 aggtttacta aaaatccgta aagaacttca attgtacgcc aacttaagac catgtaactt
     7081 tgcatccgac tctcttttag acttatctcc aatcaagcca caatttgcta aaggtactga
     7141 cttcgttgtt gtcagagaat tagtgggagg tatttacttt ggtaagagaa aggaagacga
     7201 tggtgatggt gtcgcttggg atagtgaaca atacaccgtt ccagaagtgc aaagaatcac
     7261 aagaatggcc gctttcatgg ccctacaaca tgagccacca ttgcctattt ggtccttgga
     7321 taaagctaat gttttggcct cttcaagatt atggagaaaa actgtggagg aaaccatcaa
     7381 gaacgaattc cctacattga aggttcaaca tcaattgatt gattctgccg ccatgatcct
     7441 agttaagaac ccaacccacc taaatggtat tataatcacc agcaacatgt ttggtgatat
     7501 catctccgat gaagcctccg ttatcccagg ttccttgggt ttgttgccat ctgcgtcctt
     7561 ggcctctttg ccagacaaga acaccgcatt tggtttgtac gaaccatgcc acggttctgc
     7621 tccagatttg ccaaagaata aggtcaaccc tatcgccact atcttgtctg ctgcaatgat
     7681 gttgaaattg tcattgaact tgcctgaaga aggtaaggcc attgaagatg cagttaaaaa
     7741 ggttttggat gcaggtatca gaactggtga tttaggtggt tccaacagta ccaccgaagt
     7801 cggtgatgct gtcgccgaag aagttaagaa aatccttgct taaaaagatt ctcttttttt
     7861 atgatatttg tacataaact ttataaatga aattcataat agaaacgaca cgaaattaca
     7921 aaatggaata tgttcatagg gtagacgaaa ctatatacgc aatctacata catttatcaa
     7981 gaaggagaaa aaggaggatg taaaggaata caggtaagca aattgatact aatggctcaa
     8041 cgtgataagg aaaaagaatt gcactttaac attaatattg acaaggagga gggcaccaca
     8101 caaaaagtta ggtgtaacag aaaatcatga aactatgatt cctaatttat atattggagg
     8161 attttctcta aaaaaaaaaa aatacaacaa ataaaaaaca ctcaatgacc tgaccatttg
     8221 atggagttta agtcaatacc ttcttgaacc atttcccata atggtgaaag ttccctcaag
     8281 aattttactc tgtcagaaac ggccttaacg acgtagtcga tatggtgcac tctcagtaca
     8341 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
     8401 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     8461 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc
     8521 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta ggacggatcg
     8581 cttgcctgta acttacacgc gcctcgtatc ttttaatgat ggaataattt gggaatttac
     8641 tctgtgttta tttattttta tgttttgtat ttggatttta gaaa
//
