LOCUS       Exported                7045 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  harbours marker genes LEU2 and URA3.
ACCESSION   .
VERSION     .
KEYWORDS    pLU
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7045)
  AUTHORS   Michael Mulleder, Kate Campbell, Olga Matsarskaia, Florian 
            Eckerstorfer, Markus Ralser
  TITLE     Saccharomyces cerevisiae single-copy plasmids for auxotrophy 
            compensation, multiple marker selection, and for designing 
            metabolically cooperating communities
  JOURNAL   F1000Research
REFERENCE   2  (bases 1 to 7045)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7045
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     165..183
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(221..243)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             477..1280
                     /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        1658..2065
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             2066..3160
                     /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"
     primer_bind     complement(3832..3853)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4044..4063
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4280..4302
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4294..4311
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4295..4311
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4318..4337
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4318..4336
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    4345..4452
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     promoter        complement(4464..4482)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4503..4519)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4503..4519)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4516..4538)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4527..4543
                     /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)."
     promoter        complement(4551..4581)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4596..4617
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4734..4751)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4905..5493)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4985..5004)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5664..6524)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6287..6306
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6525..6629)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    6659..117
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
ORIGIN
        1 ttacatcttc ggaaaacaaa aactattttt tctttaattt ctttttttac tttctatttt
       61 taatttatat atttatatta aaaaatttaa attataatta tttttatagc acgtgatgaa
      121 aaggacccta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
      181 ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
      241 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
      301 cgtcagcggg tgttggcggc gattcggtaa tctccgaaca gaaggaagaa cgaaggaagg
      361 agcacagact tagattggta tatatacgca tatgtagtgt tgaagaaaca tgaaattgcc
      421 cagtattctt aacccaactg cacagaacaa aaacctgcag gaaacgaaga taaatcatgt
      481 cgaaagctac atataaggaa cgtgctgcta ctcatcctag tcctgttgct gccaagctat
      541 ttaatatcat gcacgaaaag caaacaaact tgtgtgcttc attggatgtt cgtaccacca
      601 aggaattact ggagttagtt gaagcattag gtcccaaaat ttgtttacta aaaacacatg
      661 tggatatctt gactgatttt tccatggagg gcacagttaa gccgctaaag gcattatccg
      721 ccaagtacaa ttttttactc ttcgaagaca gaaaatttgc tgacattggt aatacagtca
      781 aattgcagta ctctgcgggt gtatacagaa tagcagaatg ggcagacatt acgaatgcac
      841 acggtgtggt gggcccaggt attgttagcg gtttgaagca ggcggcagaa gaagtaacaa
      901 aggaacctag aggccttttg atgttagcag aattgtcatg caagggctcc ctatctactg
      961 gagaatatac taagggtact gttgacattg cgaagagcga caaagatttt gttatcggct
     1021 ttattgctca aagagacatg ggtggaagag atgaaggtta cgattggttg attatgacac
     1081 ccggtgtggg tttagatgac aagggagacg cattgggtca acagtataga accgtggatg
     1141 atgtggtctc tacaggatct gacattatta ttgttggaag aggactattt gcaaagggaa
     1201 gggatgctaa ggtagagggt gaacgttaca gaaaagcagg ctgggaagca tatttgagaa
     1261 gatgcggcca gcaaaactaa aaaactgtat tataagtaaa tgcatgtata ctaaactcac
     1321 aaattagagc ttcaatttaa ttatatcagt tattacccta tgcggtgtga aataccgcac
     1381 agatgcgtaa ggagaaaata ccgcatcagg cgcgccatcg aggagaactt ctagtatatc
     1441 tacataccta atattattgc cttattaaaa atggaatccc aacaattaca tcaaaatcca
     1501 cattctcttc aaaatcaatt gtcctgtact tccttgttca tgtgtgttca aaaacgttat
     1561 atttatagga taattatact ctatttctca acaagtaatt ggttgtttgg ccgagcggtc
     1621 taaggcgcct gattcaagaa atatcttgac cgcagttaac tgtgggaata ctcaggtatc
     1681 gtaagatgca agagttcgaa tctcttagca accattattt ttttcctcaa cataacgaga
     1741 acacacaggg gcgctatcgc acagaatcaa attcgatgac tggaaatttt ttgttaattt
     1801 cagaggtcgc ctgacgcata tacctttttc aactgaaaaa ttgggagaaa aaggaaaggt
     1861 gagagcgccg gaaccggctt ttcatataga atagagaagc gttcatgact aaatgcttgc
     1921 atcacaatac ttgaagttga caatattatt taaggaccta ttgttttttc caataggtgg
     1981 ttagcaatcg tcttactttc taacttttct taccttttac atttcagcaa tatatatata
     2041 tatatttcaa ggatatacca ttctaatgtc tgcccctaag aagatcgtcg ttttgccagg
     2101 tgaccacgtt ggtcaagaaa tcacagccga agccattaag gttcttaaag ctatttctga
     2161 tgttcgttcc aatgtcaagt tcgatttcga aaatcattta attggtggtg ctgctatcga
     2221 tgctacaggt gttccacttc cagatgaggc gctggaagcc tccaagaagg ctgatgccgt
     2281 tttgttaggt gctgtgggtg gtcctaaatg gggtaccggt agtgttagac ctgaacaagg
     2341 tttactaaaa atccgtaaag aacttcaatt gtacgccaac ttaagaccat gtaactttgc
     2401 atccgactct cttttagact tatctccaat caagccacaa tttgctaaag gtactgactt
     2461 cgttgttgtc agagaattag tgggaggtat ttactttggt aagagaaagg aagacgatgg
     2521 tgatggtgtc gcttgggata gtgaacaata caccgttcca gaagtgcaaa gaatcacaag
     2581 aatggccgct ttcatggccc tacaacatga gccaccattg cctatttggt ccttggataa
     2641 agctaatgtt ttggcctctt caagattatg gagaaaaact gtggaggaaa ccatcaagaa
     2701 cgaattccct acattgaagg ttcaacatca attgattgat tctgccgcca tgatcctagt
     2761 taagaaccca acccacctaa atggtattat aatcaccagc aacatgtttg gtgatatcat
     2821 ctccgatgaa gcctccgtta tcccaggttc cttgggtttg ttgccatctg cgtccttggc
     2881 ctctttgcca gacaagaaca ccgcatttgg tttgtacgaa ccatgccacg gttctgctcc
     2941 agatttgcca aagaataagg tcaaccctat cgccactatc ttgtctgctg caatgatgtt
     3001 gaaattgtca ttgaacttgc ctgaagaagg taaggccatt gaagatgcag ttaaaaaggt
     3061 tttggatgca ggtatcagaa ctggtgattt aggtggttcc aacagtacca ccgaagtcgg
     3121 tgatgctgtc gccgaagaag ttaagaaaat ccttgcttaa aaagattctc tttttttatg
     3181 atatttgtac ataaacttta taaatgaaat tcataataga aacgacacga aattacaaaa
     3241 tggaatatgt tcatagggta gacgaaacta tatacgcaat ctacatacat ttatcaagaa
     3301 ggagaaaaag gaggatgtaa aggaatacag gtaagcaaat tgatactaat ggctcaacgt
     3361 gataaggaaa aagaattgca ctttaacatt aatattgaca aggaggaggg caccacacaa
     3421 aaagttaggt gtaacagaaa atcatgaaac tatgattcct aatttatata ttggaggatt
     3481 ttctctaaaa aaaaaaaaat acaacaaata aaaaacactc aatgacctga ccatttgatg
     3541 gagtttaagt caataccttc ttgaaccatt tcccataatg gtgaaagttc cctcaagaat
     3601 tttactctgt cagaaacggc cttacgacgt agtcgaggat cctatgcggt gtgaaatacc
     3661 gcacagatgc gtaaggagaa aataccgcat caggaaattg taagcgttaa tattttgtta
     3721 aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc
     3781 aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg
     3841 aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat
     3901 cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc
     3961 cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag
     4021 ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg
     4081 gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc
     4141 gctacagggc gcgtccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg
     4201 cgggcctctt cgctattacg ccagctggcg aaagggggat gtgctgcaag gcgattaagt
     4261 tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa cgacggccag tgaattgtaa
     4321 tacgactcac tatagggcga attggagctc caccgcggtg gcggccgctc tagaactagt
     4381 gggtcccccg ggctgcagga attcgatatc aagcttatcg ataccgtcga cctggagggg
     4441 gggcccggta ccagcttttg ttccctttag tgagggttaa tttcgagctt ggcgtaatca
     4501 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     4561 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     4621 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     4681 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     4741 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     4801 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     4861 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4921 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4981 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     5041 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     5101 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     5161 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     5221 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     5281 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     5341 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     5401 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     5461 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     5521 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     5581 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     5641 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     5701 atctgtctat ttcgttcatc catagttgcc tgactgcccg tcgtgtagat aactacgata
     5761 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     5821 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5881 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5941 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     6001 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     6061 tcccccatgt tgtgaaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     6121 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     6181 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     6241 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     6301 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     6361 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     6421 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     6481 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     6541 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     6601 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgga
     6661 tcgcttgcct gtaacttaca cgcgcctcgt atcttttaat gatggaataa tttgggaatt
     6721 tactctgtat ttatttattt ttatgttttg tatttggatt ttagaaagta aataaagaag
     6781 gtagaagagt tacggaatga agaaaaaaaa ataaacaaag gtttaaaaaa tttcaacaaa
     6841 aagcgtactt tacatatata tttattagac aagaaaagca gattaaatag atatacattc
     6901 gattaacgat aagtaaaatg taaaatcaca ggattttcgt gtgtggtctt ctacacagac
     6961 aagatgaaac aattcggcat taatacctga gagcaggaag agcaagataa aaggtagtat
     7021 ttgttggcga tccccctaga gtctt
//
