LOCUS       sequence-180663-        4075 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Universal C-terminal GFP11 tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4075)
  AUTHORS   Ruan L, Zhou C, Jin E, Kucharavy A, Zhang Y, Wen Z, Florens L, Li R
  TITLE     Cytosolic proteostasis through importing of misfolded proteins into 
            mitochondria.
  JOURNAL   Nature. 2017 Mar 16;543(7645):443-446. doi: 10.1038/nature21695. 
            Epub 2017 Mar 1.
  PUBMED    28241148
REFERENCE   2  (bases 1 to 4075)
  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: 
            "10.1038/nature21695"; journalName: "Nature"; date: "2017-03-16- 
            16"; volume: "543"; issue: "7645"; pages: "443-446"
COMMENT     Sequence Label: pFA6a-GFP11-His3MX
FEATURES             Location/Qualifiers
     source          1..4075
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..19
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1..18
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             92..139
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     terminator      163..350
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(218..242)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            425..1625
                     /label=HIS3MX6
                     /note="yeast selectable marker encoding the S. pombe his5 
                     gene, which corresponds to S. cerevisiae HIS3"
     promoter        425..768
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             769..1422
                     /codon_start=1
                     /gene="Schizosaccharomyces pombe his5"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=S. pombe his5
                     /note="yeast auxotrophic marker; corresponds to S. 
                     cerevisiae HIS3"
                     /translation="MGRRAFVERNTNETKISVAIALDKAPLPEESNFIDELITSKHTNQ
                     KGEQVIQVDTGIGFLDHMYHALAKHAGWSLRLYSRGDLIIDDHHTAEDTAIALGIAFKQ
                     AMSNFAGVKRFGHAYCPLDEALSRSVVDLSGRPYAVIDLGLKREKVGELSCEMIPHLLY
                     SFSVAAGITLHVTCLYGSNDHHRAESAFKSLAVAMRAATSLTGSSEVPSTKGVL"
     terminator      1428..1625
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(1729..1748)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1730..1748)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1835..1852)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2006..2594)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2086..2105)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2765..3556,3557..3625))
                     /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     3388..3407
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3626..3730)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3798..3816
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3854..3876)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3976..3995
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 atttaggtga cactatagaa cgcggccgcc agctgaagct tcgtacgctg caggtcgacg
       61 gatcggtggt ggttctggag gcggttctat gagagatcat atggttttgc atgaatatgt
      121 taatgctgct ggtattactt aaggcgcgcc acttctaaat aagcgaattt cttatgattt
      181 atgattttta ttattaaata agttataaaa aaaataagtg tatacaaatt ttaaagtgac
      241 tcttaggttt taaaacgaaa attcttattc ttgagtaact ctttcctgta ggtcaggttg
      301 ctttctcagg tatagtatga ggtcgctctt attgaccaca cctctaccgg cagatccgct
      361 agggataaca gggtaatata gatctgttta gcttgcctcg tccccgccgg gtcacccggc
      421 cagcgacatg gaggcccaga ataccctcct tgacagtctt gacgtgcgca gctcaggggc
      481 atgatgtgac tgtcgcccgt acatttagcc catacatccc catgtataat catttgcatc
      541 catacatttt gatggccgca cggcgcgaag caaaaattac ggctcctcgc tgcagacctg
      601 cgagcaggga aacgctcccc tcacagacgc gttgaattgt ccccacgccg cgcccctgta
      661 gagaaatata aaaggttagg atttgccact gaggttcttc tttcatatac ttccttttaa
      721 aatcttgcta ggatacagtt ctcacatcac atccgaacat aaacaaccat gggtaggagg
      781 gcttttgtag aaagaaatac gaacgaaacg aaaatcagcg ttgccatcgc tttggacaaa
      841 gctcccttac ctgaagagtc gaattttatt gatgaactta taacttccaa gcatacaaac
      901 caaaagggag aacaagtaat ccaagtagac acgggaattg gattcttgga tcacatgtat
      961 catgcactgg ctaaacatgc aggctggagc ttacgacttt actcaagagg tgatttaatc
     1021 atcgatgatc atcacactgc agaagatact gctattgcac ttggtattgc attcaagcag
     1081 gctatgagta actttgccgg cgttaaaaga tttggacatg cttattgtcc acttgacgaa
     1141 gctctttcta gaagcgtagt tgacttgtcg ggacggccct atgctgttat cgatttggga
     1201 ttaaagcgtg aaaaggttgg ggaattgtcc tgtgaaatga tccctcactt actatattcc
     1261 ttttcggtag cagctggaat tactttgcat gttacctgct tatatggtag taatgaccat
     1321 catcgtgctg aaagcgcttt taaatctctg gctgttgcca tgcgcgcggc tactagtctt
     1381 actggaagtt ctgaagtccc aagcacgaag ggagtgttgt aaagagtact gacaataaaa
     1441 agattcttgt tttcaagaac ttgtcatttg tatagttttt ttatattgta gttgttctat
     1501 tttaatcaaa tgttagcgtg atttatattt tttttcgcct cgacatcatc tgcccagatg
     1561 cgaagttaag tgcgcagaaa gtaatatcat gcgtcaatcg tatgtgaatg ctggtcgcta
     1621 tactgctgtc gattcgatac taacgccgcc atccagttta aacgagctcg aattcatcga
     1681 tgatatcaga tccactagtg gcctatgcgg ccgcggatct gccggtctcc ctatagtgag
     1741 tcgtattaat ttcgataagc caggttaacc tgcattaatg aatcggccaa cgcgcgggga
     1801 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     1861 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     1921 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     1981 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     2041 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     2101 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     2161 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     2221 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     2281 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     2341 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     2401 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     2461 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     2521 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     2581 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     2641 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     2701 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     2761 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     2821 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     2881 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     2941 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     3001 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     3061 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     3121 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     3181 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     3241 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     3301 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     3361 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     3421 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     3481 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     3541 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     3601 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     3661 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     3721 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca
     3781 tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg
     3841 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
     3901 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     3961 gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat atggacatat
     4021 tgtcgttaga acgcggctac aattaataca taaccttatg tatcatacac atacg
//
