LOCUS       sequence-491565-        5644 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Single-copy plasmid expressing the IQ-compete reporter 
            (GFP-TCS-Quencher-CL1)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5644)
  AUTHORS   Hoffman Y, Egeler A, Rodl S, Herrmann JM
  TITLE     The IQ-compete assay for measuring mitochondrial protein import 
            efficiencies in living yeast cells.
  JOURNAL   FEBS Lett. 2025 Oct 25. doi: 10.1002/1873-3468.70206.
  PUBMED    41137490
REFERENCE   2  (bases 1 to 5644)
  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: "FEBS Lett. 
            2025 Oct 25. doi: 10.1002/1873-3468.70206."
COMMENT     Sequence Label: IQ-compete reporter (episomal)
FEATURES             Location/Qualifiers
     source          1..5644
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        472..885
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             905..1618
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(935..963)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1558..1578
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             1625..1645
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2385..3188
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     misc_feature    3271..3774
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     CDS             complement(join(3881..4672,4673..4741))
                     /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     4504..4523
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4742..4846)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4949..5537
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5438..5457
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccctgaattc gcatctagat ggtagagcca caaacagccg gtacaagcaa cgatctccag
       61 gaccatctga atcatgcgcg gatgacacga actcacgacg gcgatcacag acattaaccc
      121 acagtacaga cactgcgaca acgtggcaat tcgtcgcaat accgtctcac tgaactggcc
      181 gataattgca gacgaacgcc ttgccaacag ggagttcttc agagacatgg aggctcaaaa
      241 cgaaattatt gacagcctag acatcaatag tcatacaaca gaaagcgacc acccaacttt
      301 ggctgataat agcgtataaa caatgcatac tttgtacgtt caaaatacaa tgcagtagat
      361 atatttatgc atattacata taatacatat cacataggaa gcaacaggcg cgttggactt
      421 ttaattttcg aggaccgcga atccttacat cacacccaat cccccacaag tgatccccca
      481 cacaccatag cttcaaaatg tttctactcc ttttttactc ttccagattt tctcggactc
      541 cgcgcatcgc cgtaccactt caaaacaccc aagcacagca tactaaattt cccctctttc
      601 ttcctctagg gtgtcgttaa ttacccgtac taaaggtttg gaaaagaaaa aagacaccgc
      661 ctcgtttctt tttcttcgtc gaaaaaggca ataaaaattt ttatcacgtt tctttttctt
      721 gaaaattttt ttttttgatt tttttctctt tcgatgacct cccattgata tttaagttaa
      781 taaacggtca tcaatttctc aagtttcagt ttcatttttc ttgttctatt acaacttttt
      841 ttacttcttg ctcattagaa agaaagcata gcaatctaat ctaagtttta attacaaaag
      901 atctatgagt aagggtgaag aacttttcac tggagttgtc ccaattcttg ttgaattaga
      961 tggtgatgtt aatgggcaca aattttctgt cagtggagag ggtgaaggtg atgcaacata
     1021 cggaaaactt acccttaaat ttatttgcac tactggaaaa ctacctgttc catggccaac
     1081 acttgtcact actttcactt atggtgttca atgcttttca agatacccag atcatatgaa
     1141 acggcatgac tttttcaaga gtgccatgcc cgaaggttat gtacaggaaa gaactatatt
     1201 tttcaaagat gacgggaact acaagacacg tgctgaagtc aagtttgaag gtgataccct
     1261 tgttaataga atcgagttaa aaggtattga ttttaaagaa gatggaaaca ttcttggaca
     1321 caaattggaa tacaactata actcacacaa tgtatacatc atggcagaca aacaaaagaa
     1381 tggaatcaaa gttaacttca aaattagaca caacattgaa gatggaagcg ttcaactagc
     1441 agaccattat caacaaaata ctccaattgg cgatggccct gtccttttac cagacaacca
     1501 ttacctgtcc acacaatctg ccctttcgaa agatcccaac gaaaagagag atcacatggt
     1561 ccttcttgag tttgtaacag ctgctgggat tacacatggc atggatgaac tatacaaagg
     1621 ttctgaaaac ttgtacttcc aatctggtgc ttgtaatgat tcttctgatc cattggttgt
     1681 tgctgcttct attattggta ttttgcattt gattttgtgg attttggata gattgggatc
     1741 catggcttgt aaaaattggt tttcttcttt gtctcatttt gttatacatt tgtaactcga
     1801 gtggcagtgc ttttaactaa gaattattag tcttttctgc ttattttttc atcatagttt
     1861 agaacacttt atattaacga atagtttatg aatctattta ggtttaaaaa ttgatacagt
     1921 tttataagtt actttttcaa agactcgtgc tgtctattgc ataatgcact ggaaggggaa
     1981 aaaaaaggtg cacacgcgtg gctttttctt gaatttgcag tttgaaaaat gctgccaatg
     2041 agacgacggg gtcatcacgg ctcatcatgc gccaaacaaa tgtgtgcaat acacgctcgg
     2101 atgactgcat gatgaccgca ctgactgggg acagcagatc cacctaagcc tgtgagagaa
     2161 gcagacaccc gacagatcaa ggcagttaac tagtgcactg cagtacacgg tttccttgaa
     2221 atttttttga ttcggtaatc tccgaacaga aggaagaacg aaggaaggag cacagactta
     2281 gattggtata tatacgcata tgtagtgttg aagaaacatg aaattgccca gtattcttaa
     2341 cccaactgca cagaacaaaa acgtgcagga aacgaagata aatcatgtcg aaagctacat
     2401 ataaggaacg tgctgctact catcctagtc ctgttgctgc caagctattt aatatcatgc
     2461 acgaaaagca aacaaacttg tgtgcttcat tggatgttcg taccaccaag gaattactgg
     2521 agttagttga agcattaggt cccaaaattt gtttactaaa aacacatgtg gatatcttga
     2581 ctgatttttc catggagggc acagttaagc cgctaaaggc attatccgcc aagtacaatt
     2641 ttttactctt cgaggacaga aaatttgctg acattggtaa tacagtcaaa ttgcagtact
     2701 ctgcgggtgt atacagaata gcagaatggg cagacattac gaatgcacac ggtgtggtgg
     2761 gcccaggtat tgttagcggt ttgaagcagg cggcagaaga agtaacaaag gaacctagag
     2821 gccttttgat gttagcagaa ttgtcatgca agggctccct atctactgga gaatatacta
     2881 agggtactgt tgacattgcg aagagcgaca aagattttgt tatcggcttt attgctcaaa
     2941 gagacatggg tggaagagat gaaggttacg attggttgat tatgacaccc ggtgtgggtt
     3001 tagatgacaa gggagatgca ttgggtcaac agtatagaac cgtggatgat gtggtttcta
     3061 caggatctga cattattatt gttggaagag gactatttgc aaagggaagg gatgctaagg
     3121 tagagggtga acgttacaga aaagcaggct gggaagcata tttgagaaga tgcggccagc
     3181 aaaactaaaa aactgtatta taagtaaatg catgtatact aaactcacaa attagagctt
     3241 caatttaatt atatcagtta ttacccgagt atcacgtgct ataaaaataa ttataattta
     3301 aattttttaa tataaatata taaattaaaa atagaaagta aaaaaagaaa ttaaagaaaa
     3361 aatagttttt gttttccgaa gatgtaaaag actctagggg gatcgccaac aaatactacc
     3421 ttttatcttg ctcttcctgc tctcaggtat taatgccgaa ttgtttcatc ttgtctgtgt
     3481 agaagaccac acacgaaaat cctgtgattt tacattttac ttatcgttaa tcgaatgtat
     3541 atctatttaa tctgcttttc ttgtctaata aatatatatg taaagtacgc tttttgttga
     3601 aattttttaa acctttgttt attttttttt cttcattccg taactcttct accttcttta
     3661 tttactttct aaaatccaaa tacaaaacat aaaaataaat aaacacagag taaattccca
     3721 aattattcca tcattaaaag atacgaggcg cgtgtaagtt acaggcaagc gatcccgagc
     3781 ggccgcgatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     3841 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     3901 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     3961 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     4021 gggacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     4081 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     4141 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     4201 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     4261 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     4321 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     4381 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     4441 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     4501 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     4561 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     4621 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     4681 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     4741 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     4801 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     4861 aagtgccacc tgtcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     4921 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     4981 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     5041 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     5101 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     5161 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     5221 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     5281 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     5341 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     5401 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     5461 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     5521 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     5581 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtgcgg
     5641 ccgc
//
