LOCUS       sequence-75757-e        7821 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7821)
  AUTHORS   Markus SM, Omer S, Baranowski K, Lee WL
  TITLE     Improved Plasmids for Fluorescent Protein Tagging of Microtubules in
            Saccharomyces cerevisiae.
  JOURNAL   Traffic. 2015 Jul;16(7):773-86. doi: 10.1111/tra.12276. Epub 2015 
            Apr 28.
  PUBMED    25711127
REFERENCE   2  (bases 1 to 7821)
  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: "10.1111/tra.12276";
            journalName: "Traffic"; date: "2015-07"; volume: "16"; issue: "7"; 
            pages: "773-86"
COMMENT     Sequence Label: pHIS3p:CloverGFP-Tub1+3'UTR::LEU2
FEATURES             Location/Qualifiers
     source          1..7821
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     72..94
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(132..150)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        218..322
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(323..391,392..1183)
                     /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(541..560)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1354..1942
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1843..1862
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2096..2113
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     2200..2219
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2200..2218
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             complement(2776..3870)
                     /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"
     promoter        complement(3871..4278)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     promoter        4641..4827
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             join(4834..4836,4837..4839,4840..5550)
                     /codon_start=1
                     /product="bright green-yellow fluorescent protein derived 
                     from GFP (Lam et al., 2012)"
                     /label=Clover
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSALSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(4879..4900)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     5487..5508
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     RBS             7035..7043
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     promoter        complement(7675..7693)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(7676..7693)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
       61 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
      121 gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
      181 ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
      241 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
      301 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
      361 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
      421 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
      481 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
      541 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
      601 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
      661 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
      721 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
      781 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
      841 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
      901 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
      961 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     1021 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     1081 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     1141 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     1201 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     1261 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     1321 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     1381 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     1441 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     1501 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     1561 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     1621 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     1681 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     1741 tgagcattga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     1801 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     1861 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     1921 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     1981 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     2041 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     2101 gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
     2161 gccgattcat taatgcaggt taacctggct tatcgaaatt aatacgactc actataggga
     2221 gaccggcaga tccgcggccg cataggccac tagttaacta tgcggtatca gagcagattg
     2281 tactgagagt gcaccatatc gactacgtcg ttaaggccgt ttctgacaga gtaaaattct
     2341 tgagggaact ttcaccatta tgggaaatgg ttcaagaagg tattgactta aactccatca
     2401 aatggtcagg tcattgagtg ttttttattt gttgtatttt ttttttttta gagaaaatcc
     2461 tccaatatat aaattaggaa tcatagtttc atgattttct gttacaccta actttttgtg
     2521 tggtgccctc ctccttgtca atattaatgt taaagtgcaa ttctttttcc ttatcacgtt
     2581 gagccattag tatcaatttg cttacctgta ttcctttaca tcctcctttt tctccttctt
     2641 gataaatgta tgtagattgc gtatatagtt tcgtctaccc tatgaacata ttccattttg
     2701 taatttcgtg tcgtttctat tatgaatttc atttataaag tttatgtaca aatatcataa
     2761 aaaaagagaa tctttttaag caaggatttt cttaacttct tcggcgacag catcaccgac
     2821 ttcggtggta ctgttggaac cacctaaatc accagttctg atacctgcat ccaaaacctt
     2881 tttaactgca tcttcaatgg ccttaccttc ttcaggcaag ttcaatgaca atttcaacat
     2941 cattgcagca gacaagatag tggcgatagg gttgacctta ttctttggca aatctggagc
     3001 agaaccgtgg catggttcgt acaaaccaaa tgcggtgttc ttgtctggca aagaggccaa
     3061 ggacgcagat ggcaacaaac ccaaggaacc tgggataacg gaggcttcat cggagatgat
     3121 atcaccaaac atgttgctgg tgattataat accatttagg tgggttgggt tcttaactag
     3181 gatcatggcg gcagaatcaa tcaattgatg ttgaaccttc aatgtaggga attcgttctt
     3241 gatggtttcc tccacagttt ttctccataa tcttgaagag gccaaaacat tagctttatc
     3301 caaggaccaa ataggcaatg gtggctcatg ttgtagggcc atgaaagcgg ccattcttgt
     3361 gattctttgc acttctggaa cggtgtattg ttcactatcc caagcgacac catcaccatc
     3421 gtcttccttt ctcttaccaa agtaaatacc tcccactaat tctctgacaa caacgaagtc
     3481 agtaccttta gcaaattgtg gcttgattgg agataagtct aaaagagagt cggatgcaaa
     3541 gttacatggt cttaagttgg cgtacaattg aagttcttta cggattttta gtaaaccttg
     3601 ttcaggtcta acactaccgg taccccattt aggaccaccc acagcaccta acaaaacggc
     3661 atcagccttc ttggaggctt ccagcgcctc atctggaagt gggacacctg tagcatcgat
     3721 agcagcacca ccaattaaat gattttcgaa atcgaacttg acattggaac gaacatcaga
     3781 aatagcttta agaaccttaa tggcttcggc tgtgatttct tgaccaacgt ggtcacctgg
     3841 caaaacgacg atcttcttag gggcagacat tagaatggta tatccttgaa atatatatat
     3901 atatattgct gaaatgtaaa aggtaagaaa agttagaaag taagacgatt gctaaccacc
     3961 tattggaaaa aacaataggt ccttaaataa tattgtcaac ttcaagtatt gtgatgcaag
     4021 catttagtca tgaacgcttc tctattctat atgaaaagcc ggttccggcg ctctcacctt
     4081 tcctttttct cccaattttt cagttgaaaa aggtatatgc gtcaggcgac ctctgaaatt
     4141 aacaaaaaat ttccagtcat cgaatttgat tctgtgcgat agcgcccctg tgtgttctcg
     4201 ttatgttgag gaaaaaaata atggttgcta agagattcga actcttgcat cttacgatac
     4261 ctgagtattc ccacagttaa ctgcggtcaa gatatttctt gaatcaggcg ccttagaccg
     4321 ctcggccaaa caaccaatta cttgttgaga aatagagtat aattatccta taaatataac
     4381 gtttttgaac acacatgaac aaggaagtac aggacaattg attttgaaga gaatgtggat
     4441 tttgatgtaa ttgttgggat tccattttta ataaggcaat aatattaggt atgtagatat
     4501 actagaagtt ctcctcgacc ggtcgatatg cggtgtgaaa taccgcacag atgcgtaagg
     4561 agaaaatacc gcatcagggg cgcgccttaa ttaaccacta aagggaacaa aagctggtac
     4621 cctattactc ttggcctcct ctagtacact ctatattttt ttatgcctcg gtaatgattt
     4681 tcattttttt tttccaccta gcggatgact cttttttttt cttagcgatt ggcattatca
     4741 cataatgaat tatacattat ataaagtaat gtgatttctt cgaagaatat actaaaaaat
     4801 gagcaggcaa gataaacgaa ggcaaagctc gagatggtga gcaagggcga ggagctgttc
     4861 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
     4921 gtccgcggcg agggcgaggg cgatgccacc aacggcaagc tgaccctgaa gttcatctgc
     4981 accaccggca agctgcccgt gccctggccc accctcgtga ccaccttcgg ctacggcgtg
     5041 gcctgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
     5101 cccgaaggct acgtccagga gcgcaccatc tctttcaagg acgacggtac ctacaagacc
     5161 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     5221 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaactt caacagccac
     5281 aacgtctata tcacggccga caagcagaag aacggcatca aggctaactt caagatccgc
     5341 cacaacgttg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     5401 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gccatcagtc cgccctgagc
     5461 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     5521 attacacatg gcatggacga gctgtacaag ggatccatga gagaagttat tagtattaat
     5581 ggtatgttcg atttgcccgt ccaggctaga tctttttttt gttagttcat tttggcgttt
     5641 catctttcat tactaacttc aacgcaaaat tttttttttt ttgatttctc tttacagtcg
     5701 gtcaagctgg ttgtcagatt ggtaatgcct gttgggaatt atattccctt gagcacggta
     5761 ttaagccgga tggacatcta gaagatggcc tttcaaagcc gaagggagga gaagagggtt
     5821 tctccacgtt tttccatgaa accggctacg gtaagttcgt tccaagggct atttacgtgg
     5881 atttagagcc caacgttatt gacgaagtcc gtaacggtcc ttacaaggac ttgttccatc
     5941 cagaacagtt gatcagtggt aaggaggacg cggctaataa ttatgcaaga ggccattaca
     6001 ccgttggtag agaaattttg ggcgatgttc tggataggat tagaaaactg gcagaccaat
     6061 gtgatgggtt acaagggttc ttgtttaccc attctcttgg tggtggtact ggttccggtc
     6121 tagggtcctt actgttggaa gaactatccg ctgaatacgg taagaaatcc aagctggaat
     6181 ttgccgtata ccctgctcca caagtgtcta cttctgtggt tgagccttac aacacagttt
     6241 taactacaca tactacattg gaacatgcag attgtacttt catggtcgat aatgaggcta
     6301 tttacgacat gtgcaaaaga aacttggata tcccaagacc aagctttgca aacttaaaca
     6361 acctaattgc tcaagtggta tcatctgtta cagcatcatt gagattcgac ggttcattaa
     6421 atgtagattt gaacgaattt caaaccaatt tggttccata tccaagaatt catttcccct
     6481 tagtttcata ttctccagtc ttatccaaat caaaggcatt ccatgagtcc aactctgtgt
     6541 cagaaattac aaacgcttgt tttgaacctg gtaaccagat ggtcaagtgt gatccaagag
     6601 atggtaaata catggctact tgtctgttat acaggggtga tgtggtaaca agagatgttc
     6661 aaagagctgt cgagcaggtg aaaaacaaga agaccgtcca attggttgat tggtgtccaa
     6721 ctggtttcaa gatcggtatt tgctacgaac ctccaactgc cacaccaaac tcacaattgg
     6781 ccactgtgga tagggccgtc tgtatgttgt caaataccac atccattgct gaggcttgga
     6841 agagaatcga tagaaaattc gatttaatgt atgccaaacg tgctttcgtc cactggtatg
     6901 tcggtgaagg tatggaagaa ggtgaattca ccgaagctag agaagatttg gctgctttag
     6961 aaagagatta catcgaagtg ggtgccgact catacgctga ggaagaggaa ttttaaggtt
     7021 cactctcgcc cccaacctcc ttatcctttc tatatttcaa gtatattgta aaaaaaaaaa
     7081 ttccccctac tgttatatct ctttggtgat atcttatcta taatctactt attatatttc
     7141 tctaatatct atgtacaaaa aataaaaaaa aaaaccattc tccttctcta acctgcattt
     7201 gatgaaagct tagtaagtct catcgtagtt gatgggccaa agacatttgt acttgtttca
     7261 tcagattctt acatcaccca tttccatgat tatttcacgg atttcattag taattgcatt
     7321 tttgtatccc tagttccttt tctcccgcct tgccgtcgaa ttgaaaaaaa aaatggaaat
     7381 gaggacatct tccattgata tccaggacaa agcaagagta gaagtaacta ttagcaagga
     7441 gaattccaaa aatagtgaac ttcaaggtaa gcagatctat agtttcaata tctgattttc
     7501 tgcttttcaa aacttaaaat tcattagaag gtaaaacaag aacccgtaaa attaaagtta
     7561 ccgatattat caggtttttc accccatgtc tactatccca tttagaaaga actatgtgtt
     7621 caaaaactgg gccggtcgac ctgcagcgta cgaagcttca gctggcggcc gcgttctata
     7681 gtgtcaccta aatcgtatgt gtatgataca taaggttatg tattaattgt agccgcgttc
     7741 taacgacaat atgtccatat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     7801 aagccagccc cgacacccgc c
//
