LOCUS       sequence-352641-        5584 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Eleventh beta-strand of GFP (GFP11) fused to a mitochondria signal 
            sequence for mitochondrial targeting, HA tagged in yeast plasmid 
            with URA marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5584)
  AUTHORS   Cortes Sanchon A, Santhosh Kumar H, Mantovani M, Osinnii I, Mateos 
            JM, Kaech A, Shcherbakov D, Akbergenov R, Bottger EC
  TITLE     ER-misfolded proteins become sequestered with mitochondria and 
            impair mitochondrial function.
  JOURNAL   Commun Biol. 2021 Dec 2;4(1):1350. doi: 10.1038/s42003-021-02873-w.
  PUBMED    34857875
REFERENCE   2  (bases 1 to 5584)
  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: "Commun 
            Biol."; date: "2021-12-2"; volume: "4(1):1350. doi"; pages: " 
            10.1038/s42003-021-02873-w"
COMMENT     Sequence Label: p406 – TDH3-MTS-GFP11-Ura
FEATURES             Location/Qualifiers
     source          1..5584
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     128..147
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     381..398
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    515..536
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(551..556,557..574,575..581)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    589..605
                     /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     594..616
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     613..629
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     613..629
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     648..668
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        650..668
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        693..1336
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     1230..1253
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             1617..1643
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1683..1730
                     /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      1742..1989
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(1758..1776)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2007..2026)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2008..2026)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2036..2053)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2036..2052)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2045..2067)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2193..2648
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2280..2299)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2490..2511
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2779..3582)
                     /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"
     promoter        complement(3583..3798)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(3822..3841)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3941..3963
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4001..4019)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4087..4191
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4192..4260,4261..5052)
                     /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(4410..4429)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(5223..5584,1..227)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
       61 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
      121 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
      181 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
      241 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
      301 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
      361 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac
      421 gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc
      481 ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg
      541 caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat
      601 aacaatttca cacaggaaac agctatgacc atgattacgc caagcgcgca attaaccctc
      661 actaaaggga acaaaagctg gagctcagtt tatcattatc aatactcgcc atttcaaaga
      721 atacgtaaat aattaatagt agtgattttc ctaactttat ttagtcaaaa aattagcctt
      781 ttaattctgc tgtaacccgt acatgcccaa aatagggggc gggttacaca gaatatataa
      841 catcgtaggt gtctgggtga acagtttatt cctggcatcc actaaatata atggagcccg
      901 ctttttaagc tggcatccag aaaaaaaaag aatcccagca ccaaaatatt gttttcttca
      961 ccaaccatca gttcataggt ccattctctt agcgcaacta cagagaacag gggcacaaac
     1021 aggcaaaaaa cgggcacaac ctcaatggag tgatgcaacc tgcctggagt aaatgatgac
     1081 acaaggcaat tgacccacgc atgtatctat ctcattttct tacaccttct attaccttct
     1141 gctctctctg atttggaaaa agctgaaaaa aaaggttgaa accagttccc tgaaattatt
     1201 cccctacttg actaataagt atataaagac ggtaggtatt gattgtaatt ctgtaaatct
     1261 atttcttaaa cttcttaaat tctactttta tagttagtct tttttttagt tttaaaacac
     1321 cagaacttag tttcgacgga ttctagaact agtgagctca agcttcgaat tctgcagtcg
     1381 acatggcctc cactcgtgtc ctcgcctctc gcctggcctc ccggatggct gcttccgcca
     1441 aggttgcccg ccctgctgtc cgcgttgctc aggtcagcaa gcgcaccatc cagactggct
     1501 cccccctcca gaccctcaag cgcacccaga tgacctccat cgtcaacgcc accacccgcc
     1561 aggctttcca gaagcgcgcc tactcttccg gatccggtga cggtccggtc gccacctacc
     1621 catacgatgt tccagattac gctggtcgac ggatcggtgg tggttctgga ggcggttcta
     1681 tgagagatca tatggttttg catgaatatg ttaatgctgc tggtattact taagcctcga
     1741 gtcatgtaat tagttatgtc acgcttacat tcacgccctc cccccacatc cgctctaacc
     1801 gaaaaggaag gagttagaca acctgaagtc taggtcccta tttatttttt tatagttatg
     1861 ttagtattaa gaacgttatt tatatttcaa atttttcttt tttttctgta cagacgcgtg
     1921 tacgcatgta acattatact gaaaaccttg cttgagaagg ttttgggacg ctcgaaggct
     1981 ttaatttgcg gccggtaccc aattcgccct atagtgagtc gtattacgcg cgctcactgg
     2041 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg
     2101 cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt
     2161 cccaacagtt gcgcagcctg aatggcgaat ggacgcgccc tgtagcggcg cattaagcgc
     2221 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     2281 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     2341 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     2401 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     2461 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     2521 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     2581 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
     2641 tacaatttcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     2701 gggtaataac tgatataatt aaattgaagc tctaatttgt gagtttagta tacatgcatt
     2761 tacttataat acagtttttt agttttgctg gccgcatctt ctcaaatatg cttcccagcc
     2821 tgcttttctg taacgttcac cctctacctt agcatccctt ccctttgcaa atagtcctct
     2881 tccaacaata ataatgtcag atcctgtaga gaccacatca tccacggttc tatactgttg
     2941 acccaatgcg tctcccttgt catctaaacc cacaccgggt gtcataatca accaatcgta
     3001 accttcatct cttccaccca tgtctctttg agcaataaag ccgataacaa aatctttgtc
     3061 gctcttcgca atgtcaacag tacccttagt atattctcca gtagataggg agcccttgca
     3121 tgacaattct gctaacatca aaaggcctct aggttccttt gttacttctt ctgccgcctg
     3181 cttcaaaccg ctaacaatac ctgggcccac cacaccgtgt gcattcgtaa tgtctgccca
     3241 ttctgctatt ctgtatacac ccgcagagta ctgcaatttg actgtattac caatgtcagc
     3301 aaattttctg tcttcgaaga gtaaaaaatt gtacttggcg gataatgcct ttagcggctt
     3361 aactgtgccc tccatggaaa aatcagtcaa gatatccaca tgtgttttta gtaaacaaat
     3421 tttgggacct aatgcttcaa ctaactccag taattccttg gtggtacgaa catccaatga
     3481 agcacacaag tttgtttgct tttcgtgcat gatattaaat agcttggcag caacaggact
     3541 aggatgagta gcagcacgtt ccttatatgt agctttcgac atgatttatc ttcgtttcct
     3601 gcaggttttt gttctgtgca gttgggttaa gaatactggg caatttcatg tttcttcaac
     3661 actacatatg cgtatatata ccaatctaag tctgtgctcc ttccttcgtt cttccttctg
     3721 ttcggagatt accgaatcaa aaaaatttca aggaaaccga aatcaaaaaa aagaataaaa
     3781 aaaaaatgat gaattgaaaa ggtggtatgg tgcactctca gtacaatctg ctctgatgcc
     3841 gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     3901 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     3961 aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt
     4021 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga
     4081 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     4141 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     4201 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     4261 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     4321 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     4381 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     4441 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     4501 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     4561 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     4621 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     4681 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     4741 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     4801 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     4861 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     4921 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     4981 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     5041 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     5101 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     5161 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     5221 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     5281 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     5341 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     5401 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     5461 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     5521 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     5581 acct
//
