LOCUS       Exported                6518 bp DNA     circular SYN 14-NOV-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 6518)
  AUTHORS   The Francis Crick Institute
  TITLE     Direct Submission
  JOURNAL   Exported Sep 8, 2021 from SnapGene Viewer 5.3.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6518
                     /organism="recombinant plasmid"
                     /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"
     primer_bind     343..362
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        507..694
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             695..1357
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     primer_bind     complement(1753..1774)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1965..1984
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2201..2223
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2215..2232
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2216..2232
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2239..2258
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2239..2257
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2260..2309
                     /label=p16NotI_GAP_f
     misc_feature    2266..2283
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     2283..2309
                     /label=p16_GAPprom_f
     promoter        2284..2950
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase"
     primer_bind     2284..2305
                     /label=NotI_GAPprom_f
     primer_bind     2794..2813
                     /label=GPD fw 1
     primer_bind     2841..2860
                     /label=GPD fw 2
     primer_bind     2868..2891
                     /label=p426-sense(GPD_pr)
     primer_bind     complement(2926..2956)
                     /label=mCherry_GAPprom_r
     primer_bind     complement(2951..2963)
                     /label=XhoI_GAPprom_r
     gene            2975..3691
                     /product="Yeast codon optimised (Pombe) enhanced cyan 
                     fluorescent protein (mEx: 435 nm, mEm: 485 nm). From Watson
                     et al, Gene 2008). Complete sequence homology to pAW8-yECFP
                     (http://www.euroscarf.de/plasmid_details.php?accno=P30547) 
                     "
                     /label=eCFP
     primer_bind     3704..3732
                     /label=ADH1term_mCherry_r
     terminator      3712..3899
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     3712..3727
                     /label=NheI_ADHIterm_f
     primer_bind     complement(3875..3924)
                     /label=p16SalI_ADH1_r
     primer_bind     complement(3883..3947)
                     /label=p16_ADH1term_r
     primer_bind     complement(3886..3908)
                     /label=SalI_ADH1term_r
     misc_feature    3904..3925
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     3921..3947
                     /label=uni_p16_f
     primer_bind     3921..3947
                     /label=ADH1term_p16_f
     promoter        complement(3937..3955)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3937..3955)
                     /label=T3
                     /note="T3 promoter, forward primer"
     primer_bind     complement(3976..3992)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3976..3992)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3989..4011)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4000..4016
                     /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(4024..4054)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4069..4090
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4207..4224)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4378..4966)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4458..4477)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5137..5997)
                     /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     5760..5779
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5998..6102)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    join(6132..6518,1..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 tgttggcgga cgtcggggct ggcttaacta tgcggcatca gagcagattg
      361 tactgagagt gcaccataat tccgttttaa gagcttggtg agcgctagga gtcactgcca
      421 ggtatcgttt gaacacggca ttagtcaggg aagtcataac acagtccttt cccgcaattt
      481 tctttttcta ttactcttgg cctcctctag tacactctat atttttttat gcctcggtaa
      541 tgattttcat tttttttttt ccacctagcg gatgactctt tttttttctt agcgattggc
      601 attatcacat aatgaattat acattatata aagtaatgtg atttcttcga agaatatact
      661 aaaaaatgag caggcaagat aaacgaaggc aaagatgaca gagcagaaag ccctagtaaa
      721 gcgtattaca aatgaaacca agattcagat tgcgatctct ttaaagggtg gtcccctagc
      781 gatagagcac tcgatcttcc cagaaaaaga ggcagaagca gtagcagaac aggccacaca
      841 atcgcaagtg attaacgtcc acacaggtat agggtttctg gaccatatga tacatgctct
      901 ggccaagcat tccggctggt cgctaatcgt tgagtgcatt ggtgacttac acatagacga
      961 ccatcacacc actgaagact gcgggattgc tctcggtcaa gcttttaaag aggccctagg
     1021 ggccgtgcgt ggagtaaaaa ggtttggatc aggatttgcg cctttggatg aggcactttc
     1081 cagagcggtg gtagatcttt cgaacaggcc gtacgcagtt gtcgaacttg gtttgcaaag
     1141 ggagaaagta ggagatctct cttgcgagat gatcccgcat tttcttgaaa gctttgcaga
     1201 ggctagcaga attaccctcc acgttgattg tctgcgaggc aagaatgatc atcaccgtag
     1261 tgagagtgcg ttcaaggctc ttgcggttgc cataagagaa gccacctcgc ccaatggtac
     1321 caacgatgtt ccctccacca aaggtgttct tatgtagtga caccgattat ttaaagctgc
     1381 agcatacgat atatatacat gtgtatatat gtatacctat gaatgtcagt aagtatgtat
     1441 acgaacagta tgatactgaa gatgacaagg taatgcatca ttctatacgt gtcattctga
     1501 acgaggcgcg ctttcctttt ttctttttgc tttttctttt tttttctctt gaactcgaga
     1561 tcctatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggaaatt
     1621 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
     1681 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     1741 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
     1801 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
     1861 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
     1921 tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa
     1981 ggagcgggcg ctagggcgct ggctggcaag tgtagcggtc acgctgcgcg taaccaccac
     2041 acccgccgcg cttaatgcgc cgctacaggg cgcgtccatt cgccattcag gctgcgcaac
     2101 tgttgggaag ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga
     2161 tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa
     2221 acgacggcca gtgaattgta atacgactca ctatagggcg aattggagct ccaccgcggt
     2281 ggctcattat caatactgcc atttcaaaga atacgtaaat aattaatagt agtgattttc
     2341 ctaactttat ttagtcaaaa aattagcctt ttaattctgc tgtaacccgt acatgcccaa
     2401 aatagggggc gggttacaca gaatatataa catcgtaggt gtctgggtga acagtttatt
     2461 cctggcatcc actaaatata atggagcccg ctttttaagc tggcatccag aaaaaaaaag
     2521 aatcccagca ccaaaatatt gttttcttca ccaaccatca gttcataggt ccattctctt
     2581 agcgcaacta cagagaacag gggcacaaac aggcaaaaaa cgggcacaac ctcaatggag
     2641 tgatgcaacc tgcctggagt aaatgatgac acaaggcaat tgacccacgc atgtatctat
     2701 ctcattttct tacaccttct attaccttct gctctctctg atttggaaaa agctgaaaaa
     2761 aaaggttgaa accagttccc tgaaattatt cccctacttg actaataagt atataaagac
     2821 ggtaggtatt gattgtaatt ctgtaaatct atttcttaaa cttcttaaat tctactttta
     2881 tagttagtct tttttttagt tttaaaacac caagaactta gtttcgaata aacacacata
     2941 aacaaacaaa ggtgacggtg ctggtttaat taacatgtct aaaggtgaag aattattcac
     3001 tggtgttgtc ccaattttgg ttgaattaga tggtgatgtt aatggtcaca aattttctgt
     3061 ctccggtgaa ggtgaaggtg atgctactta cggtaaattg accttaaaat ttatttgtac
     3121 tactggtaaa ttgccagttc catggccaac cttagtcact actttaactt ggggtgttca
     3181 atgtttttct agatacccag atcatatgaa acaacatgac tttttcaagt ctgccatgcc
     3241 agaaggttat gttcaagaaa gaactatttt tttcaaagat gacggtaact acaagaccag
     3301 agctgaagtc aagtttgaag gtgatacctt agttaataga atcgaattaa aaggtattga
     3361 ttttaaagaa gatggtaaca ttttaggtca caaattggaa tacatttata actctcacaa
     3421 tgtttacatc actgctgaca aacaaaagaa tggtatcaaa gctaacttca aaattagaca
     3481 caacattgaa gatggttctg ttcaattagc tgaccattat caacaaaata ctccaattgg
     3541 tgatggtcca gtcttgttac cagacaacca ttacttatcc actcaatctg ccttatccaa
     3601 agatccaaac gaaaagagag accacatggt cttgttagaa tttgttactg ctgctggtat
     3661 tacccatggt atggatgaat tgtacaaata aggcgcgcca cttctaaata agcgaatttc
     3721 ttatgattta tgatttttat tattaaataa gttataaaaa aaataagtgt atacaaattt
     3781 taaagtgact cttaggtttt aaaacgaaaa ttcttattct tgagtaactc tttcctgtag
     3841 gtcaggttgc tttctcaggt atagtatgag gtcgctctta ttgaccacac ctctaccggt
     3901 cgacctggag ggggggcccg gtaccagctt ttgttccctt tagtgagggt taatttcgag
     3961 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     4021 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     4081 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     4141 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     4201 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     4261 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     4321 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     4381 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     4441 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     4501 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     4561 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     4621 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     4681 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     4741 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     4801 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4861 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4921 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4981 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     5041 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     5101 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     5161 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactgc ccgtcgtgta
     5221 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     5281 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     5341 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     5401 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     5461 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     5521 gcgagttaca tgatccccca tgttgtgaaa aaaagcggtt agctccttcg gtcctccgat
     5581 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     5641 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     5701 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     5761 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     5821 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     5881 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     5941 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     6001 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     6061 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6121 gccacctgac ggatcgcttg cctgtaactt acacgcgcct cgtatctttt aatgatggaa
     6181 taatttggga atttactctg tatttattta tttttatgtt ttgtatttgg attttagaaa
     6241 gtaaataaag aaggtagaag agttacggaa tgaagaaaaa aaaataaaca aaggtttaaa
     6301 aaatttcaac aaaaagcgta ctttacatat atatttatta gacaagaaaa gcagattaaa
     6361 tagatataca ttcgattaac gataagtaaa atgtaaaatc acaggatttt cgtgtgtggt
     6421 cttctacaca gacaagatga aacaattcgg cattaatacc tgagagcagg aagagcaaga
     6481 taaaaggtag tatttgttgg cgatccccct agagtctt
//
