LOCUS       sequence-332098-        7784 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Lifeact-mTq2 to visualize actin in yeast.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7784)
  TITLE     Teusink unpublished plasmids
REFERENCE   2  (bases 1 to 7784)
  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"
COMMENT     Sequence Label: LifeAct-mTq2 pDRF1-GW
FEATURES             Location/Qualifiers
     source          1..7784
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    60..84
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             133..183
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
     regulatory      199..208
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(205..207,208..210,211..924)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Goedhart et 
                     al., 2012)"
                     /label=mTurquoise2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYFSDNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(250..271)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(511..530)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     858..879
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(934..958)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(1020..1039)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1021..1039)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      1187..1374
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1242..1266)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(1386..1403)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1386..1402)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1395..1417)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1838..3184
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     CDS             complement(3295..4098)
                     /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(4099..4314)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4339..4357)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4425..4529
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4530..4598,4599..5390)
                     /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(4748..4767)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5561..6149
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6050..6069
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6303..6320
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6437..6458
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6473..6478,6479..6496,6497..6503)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6511..6527
                     /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     6516..6538
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6535..6551
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6535..6551
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     rep_origin      complement(6581..7036)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6718..6739)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6930..6949
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 acattcaaaa gaaagaaaaa aaatataccc cagcctcgac tagtgaattc gcggccatca
       61 caagtttgta caaaaaagca ggctgctagc gctaccggac tcagatctcg agctcaagct
      121 tcgaattcca ccatgggcgt ggccgacctg atcaagaagt tcgagagcat cagcaaggag
      181 gagggggatc caccggtcgc caccatggtg agcaagggcg aggagctgtt caccggggtg
      241 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc
      301 gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc
      361 aagctgcccg tgccctggcc caccctcgtg accaccctgt cctggggcgt gcagtgcttc
      421 gcccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
      481 tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag
      541 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
      601 gaggacggca acatcctggg gcacaagctg gagtacaact actttagcga caacgtctat
      661 atcaccgccg acaagcagaa gaacggcatc aaggccaact tcaagatccg ccacaacatc
      721 gaggacggcg gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc
      781 cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccaagctgag caaagacccc
      841 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
      901 ggcatggacg agctgtacaa gtaaagcggc cgcacccagc tttcttgtac aaagtggtgt
      961 agttaattca tgatggccgc tgcaggtcga cctcgagggg gggcccggta cccaattcgc
     1021 cctatagtga gtcgtattac gcgcggatcc agctttggac ttcttcgcca gaggtttggt
     1081 caagtctcca atcaaggttg tcggcttgtc taccttgcca gaaatttacg aaaagatgga
     1141 aaagggtcaa atcgttggta gatacgttgt tgacacttct aaataagcga atttcttatg
     1201 atttatgatt tttattatta aataagttat aaaaaaaata agtgtataca aattttaaag
     1261 tgactcttag gttttaaaac gaaaattctt attcttgagt aactctttcc tgtaggtcag
     1321 gttgctttct caggtatagc atgaggtcgc tcttattgac cacacctcta ccggcatgcc
     1381 aattcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt
     1441 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc
     1501 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat ggcgcctgat gcggtatttt
     1561 ctccttacgc atctgtgcgg tatttcacac cgcataatcg gatcgtactt gttacccatc
     1621 attgaatttt gaacatccga acctgggagt tttccctgaa acagatagta tatttgaacc
     1681 tgtataataa tatatagtct agcgctttac ggaagacaat gtatgtattt cggttcctgg
     1741 agaaactatt gcatctattg cataggtaat cttgcacgtc gcatccccgg ttcattttct
     1801 gcgtttccat cttgcacttc aatagcatat ctttgttaac gaagcatctg tgcttcattt
     1861 tgtagaacaa aaatgcaacg cgagagcgct aatttttcaa acaaagaatc tgagctgcat
     1921 ttttacagaa cagaaatgca acgcgaaagc gctattttac caacgaagaa tctgtgcttc
     1981 atttttgtaa aacaaaaatg caacgcgaga gcgctaattt ttcaaacaaa gaatctgagc
     2041 tgcattttta cagaacagaa atgcaacgcg agagcgctat tttaccaaca aagaatctat
     2101 acttcttttt tgttctacaa aaatgcatcc cgagagcgct atttttctaa caaagcatct
     2161 tagattactt tttttctcct ttgtgcgctc tataatgcag tctcttgata actttttgca
     2221 ctgtaggtcc gttaaggtta gaagaaggct actttggtgt ctattttctc ttccataaaa
     2281 aaagcctgac tccacttccc gcgtttactg attactagcg aagctgcggg tgcatttttt
     2341 caagataaag gcatccccga ttatattcta taccgatgtg gattgcgcat actttgtgaa
     2401 cagaaagtga tagcgttgat gattcttcat tggtcagaaa attatgaacg gtttcttcta
     2461 ttttgtctct atatactacg tataggaaat gtttacattt tcgtattgtt ttcgattcac
     2521 tctatgaata gttcttacta caattttttt gtctaaagag taatactaga gataaacata
     2581 aaaaatgtag aggtcgagtt tagatgcaag ttcaaggagc gaaaggtgga tgggtaggtt
     2641 atatagggat atagcacaga gatatatagc aaagagatac ttttgagcaa tgtttgtgga
     2701 agcggtattc gcaatatttt agtagctcgt tacagtccgg tgcgtttttg gttttttgaa
     2761 agtgcgtctt cagagcgctt ttggttttca aaagcgctct gaagttccta tactttctag
     2821 ctagagaata ggaacttcgg aataggaact tcaaagcgtt tccgaaaacg agcgcttccg
     2881 aaaatgcaac gcgagctgcg cacatacagc tcactgttca cgtcgcacct atatctgcgt
     2941 gttgcctgta tatatatata catgagaaga acggcatagt gcgtgtttat gcttaaatgc
     3001 gtacttatat gcgtctattt atgtaggatg aaaggtagtc tagtacctcc tgtgatatta
     3061 tcccattcca tgcggggtat cgtatgcttc cttcagcact accctttagc tgttctatat
     3121 gctgccactc ctcaattgga ttagtctcat ccttcaatgc tatcatttcc tttgatattg
     3181 gatcgatccg atgataagct gtcaaacatg agaattgggt aataactgat ataattaaat
     3241 tgaagctcta atttgtgagt ttagtataca tgcatttact tataatacag ttttttagtt
     3301 ttgctggccg catcttctca aatatgcttc ccagcctgct tttctgtaac gttcaccctc
     3361 taccttagca tcccttccct ttgcaaatag tcctcttcca acaataataa tgtcagatcc
     3421 tgtagagacc acatcatcca cggttctata ctgttgaccc aatgcgtctc ccttgtcatc
     3481 taaacccaca ccgggtgtca taatcaacca atcgtaacct tcatctcttc cacccatgtc
     3541 tctttgagca ataaagccga taacaaaatc tttgtcgctc ttcgcaatgt caacagtacc
     3601 cttagtatat tctccagtag atagggagcc cttgcatgac aattctgcta acatcaaaag
     3661 gcctctaggt tcctttgtta cttcttctgc cgcctgcttc aaaccgctaa caatacctgg
     3721 gcccaccaca ccgtgtgcat tcgtaatgtc tgcccattct gctattctgt atacacccgc
     3781 agagtactgc aatttgactg tattaccaat gtcagcaaat tttctgtctt cgaagagtaa
     3841 aaaattgtac ttggcggata atgcctttag cggcttaact gtgccctcca tggaaaaatc
     3901 agtcaagata tccacatgtg tttttagtaa acaaattttg ggacctaatg cttcaactaa
     3961 ctccagtaat tccttggtgg tacgaacatc caatgaagca cacaagtttg tttgcttttc
     4021 gtgcatgata ttaaatagct tggcagcaac aggactagga tgagtagcag cacgttcctt
     4081 atatgtagct ttcgacatga tttatcttcg tttcctgcat gtttttgttc tgtgcagttg
     4141 ggttaagaat actgggcaat ttcatgtttc ttcaacacta catatgcgta tatataccaa
     4201 tctaagtctg tgctccttcc ttcgttcttc cttctgttcg gagattaccg aatcaaaaaa
     4261 atttcaagga aaccgaaatc aaaaaaaaga ataaaaaaaa aatgatgaat tgaaaagcta
     4321 attcttgaag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     4381 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     4441 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     4501 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     4561 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     4621 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     4681 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     4741 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     4801 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     4861 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     4921 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     4981 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     5041 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     5101 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     5161 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     5221 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     5281 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     5341 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     5401 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     5461 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     5521 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     5581 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     5641 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     5701 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     5761 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     5821 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     5881 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     5941 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     6001 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     6061 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     6121 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     6181 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     6241 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     6301 gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg
     6361 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     6421 gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact
     6481 ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
     6541 acagctatga ccatgattac gccaagctta ccgcatcagg aaattgtaag cgttaatatt
     6601 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     6661 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     6721 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     6781 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     6841 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     6901 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     6961 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     7021 ccgctacagg gcgcgtccat tcgccaagct tcctgaaacg gagaaacata aacaggcatt
     7081 gctgggatca cccatacatc actctgtttt gcctgacctt ttccggtaat ttgaaaacaa
     7141 acccggtctc gaagcggaga tccggcgata attaccgcag aaataaaccc atacacgaga
     7201 cgtagaacca gccgcacatg gccggagaaa ctcctgcgag aatttcgtaa actcgcgcgc
     7261 attgcatctg tatttcctaa tgcggcactt ccaggcctcg atcgagaccg tttatccatt
     7321 gcttttttgt tgtctttttc cctcgttcac agaaagtctg aagaagctat agtagaacta
     7381 tgagcttttt ttgtttctgt tttccttttt ttttttttta cctctgtgga aattgttact
     7441 ctcacactct ttagttcgtt tgtttgtttt gtttattcca attatgaccg gtgacgaaac
     7501 gtggtcgatg gtgggtaccg cttatgctcc cctccattag tttcgattat ataaaaaggc
     7561 caaatattgt attattttca aatgtcctat cattatcgtc taacatctaa tttctcttaa
     7621 attttttctc tttctttcct ataacaccaa tagtgaaaat ctttttttct tctatatcta
     7681 caaaaacttt ttttttctat caacctcgtt gataaatttt ttctttaaca atcgttaata
     7741 attaattaat tggaaaataa ccattttttc tctcttttat acac
//
