LOCUS       Exported                5582 bp ds-DNA     circular SYN 25-JUL-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5582)
  AUTHORS   Trial User
  TITLE     Direct Submission
  JOURNAL   Exported Wednesday, Nov 25, 2020 from SnapGene Viewer 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5582
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          1657..1859
                     /organism="Saccharomyces cerevisiae S288c"
                     /chromosome="I"
                     /strain="S288c"
                     /mol_type="genomic DNA"
                     /label=R64-2-1
                     /note="R64-2-1"
                     /db_xref="taxon:559292"
     promoter        317..503
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             504..1163
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALLARGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHFL
                     ESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     misc_feature    1606..1625
                     /label=sgRNA-seq2
     primer_bind     1648..1678
                     /label=tRNA_Pro-F
     promoter        1657..1859
                     /label=tRNA_Pro promoter
     gene            1757..1859
                     /gene="TRN1"
                     /locus_tag="tP(UGG)A"
                     /label=TRN1
     primer_bind     complement(1842..1900)
                     /label=tRNA_Pro-R
     misc_RNA        1876..1951
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      1963..1982
                     /label=SUP4 terminator
                     /note="transcription terminator for the S. cerevisiae SUP4 
                     tRNA gene"
     primer_bind     complement(2020..2036)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2032..2055)
                     /label=M13R-48
     protein_bind    2044..2060
                     /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(2068..2098)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2113..2134
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(2422..3010)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3181..4041)
                     /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"
     promoter        complement(4042..4146)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4173..5515
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(5136..5183)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataaatt cccgttttaa gagcttggtg agcgctagga gtcactgcca ggtatcgttt
      241 gaacacggca ttagtcaggg aagtcataac acagtccttt cccgcaattt tctttttcta
      301 ttactcttgg cctcctctag tacactctat atttttttat gcctcggtaa tgattttcat
      361 tttttttttt cccctagcgg atgactcttt ttttttctta gcgattggca ttatcacata
      421 atgaattata cattatataa agtaatgtga tttcttcgaa gaatatacta aaaaatgagc
      481 aggcaagata aacgaaggca aagatgacag agcagaaagc cctagtaaag cgtattacaa
      541 atgaaaccaa gattcagatt gcgatctctt taaagggtgg tcccctagcg atagagcact
      601 cgatcttccc agaaaaagag gcagaagcag tagcagaaca ggccacacaa tcgcaagtga
      661 ttaacgtcca cacaggtata gggtttctgg accatatgat acatgctctg gccaagcatt
      721 ccggctggtc gctaatcgtt gagtgcattg gtgacttaca catagacgac catcacacca
      781 ctgaagactg cgggattgct ctcggtcaag cttttaaaga ggccctactg gcgcgtggag
      841 taaaaaggtt tggatcagga tttgcgcctt tggatgaggc actttccaga gcggtggtag
      901 atctttcgaa caggccgtac gcagttgtcg aacttggttt gcaaagggag aaagtaggag
      961 atctctcttg cgagatgatc ccgcattttc ttgaaagctt tgcagaggct agcagaatta
     1021 ccctccacgt tgattgtctg cgaggcaaga atgatcatca ccgtagtgag agtgcgttca
     1081 aggctcttgc ggttgccata agagaagcca cctcgcccaa tggtaccaac gatgttccct
     1141 ccaccaaagg tgttcttatg tagtgacacc gattatttaa agctgcagca tacgatatat
     1201 atacatgtgt atatatgtat acctatgaat gtcagtaagt atgtatacga acagtatgat
     1261 actgaagatg acaaggtaat gcatcattct atacgtgtca ttctgaacga ggcgcgcttt
     1321 ccttttttct ttttgctttt tctttttttt tctcttgaac tcgacggatc tatgcggtgt
     1381 gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggaaattgta aacgttaata
     1441 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     1501 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     1561 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     1621 ccgtctatca gggcgatggc ccactacgtg ctcgagtccg tagtctaaat gagttacgta
     1681 cgaaagggag ccaaaagcaa tctgaccaat ttgtatatat atacatctat ccgaaaagga
     1741 agacatcaat tagtacgggc gtgtggtcta gtggtatgat tctcgctttg ggcgacttcc
     1801 tgattaaaca ggaagacaaa gcatgcgaga ggccctgggt tcaattccca gctcgccccg
     1861 gagaccgagg tctccgtttt agagctagaa atagcaagtt aaaataaggc tagtccgtta
     1921 tcaacttgaa aaagtggcac cgagtcggtg cttttttgat cctttttttg ttttttatgt
     1981 ctgagctccc tgcaggcatg caagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     2041 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     2101 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     2161 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     2221 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     2281 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     2341 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     2401 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     2461 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     2521 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     2581 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     2641 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     2701 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     2761 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     2821 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     2881 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     2941 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     3001 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     3061 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     3121 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     3181 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     3241 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     3301 cagtgctgca atgataccgc ggctcccacg ctcaccggct ccagatttat cagcaataaa
     3361 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     3421 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     3481 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     3541 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     3601 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     3661 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     3721 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     3781 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     3841 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     3901 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     3961 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     4021 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     4081 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     4141 tccgcgcaca tttccccgaa aagtgccacc tgaacgaagc atctgtgctt cattttgtag
     4201 aacaaaaatg caacgcgaga gcgctaattt ttcaaacaaa gaatctgagc tgcattttta
     4261 cagaacagaa atgcaacgcg aaagcgctat tttaccaacg aagaatctgt gcttcatttt
     4321 tgtaaaacaa aaatgcaacg cgagagcgct aatttttcaa acaaagaatc tgagctgcat
     4381 ttttacagaa cagaaatgca acgcgagagc gctattttac caacaaagaa tctatacttc
     4441 ttttttgttc tacaaaaatg catcccgaga gcgctatttt tctaacaaag catcttagat
     4501 tacttttttt ctcctttgtg cgctctataa tgcagtctct tgataacttt ttgcactgta
     4561 ggtccgttaa ggttagaaga aggctacttt ggtgtctatt ttctcttcca taaaaaaagc
     4621 ctgactccac ttcccgcgtt tactgattac tagcgaagct gcgggtgcat tttttcaaga
     4681 taaaggcatc cccgattata ttctataccg atgtggattg cgcatacttt gtgaacagaa
     4741 agtgatagcg ttgatgattc ttcattggtc agaaaattat gaacggtttc ttctattttg
     4801 tctctatata ctacgtatag gaaatgttta cattttcgta ttgttttcga ttcactctat
     4861 gaatagttct tactacaatt tttttgtcta aagagtaata ctagagataa acataaaaaa
     4921 tgtagaggtc gagtttagat gcaagttcaa ggagcgaaag gtggatgggt aggttatata
     4981 gggatatagc acagagatat atagcaaaga gatacttttg agcaatgttt gtggaagcgg
     5041 tattcgcaat attttagtag ctcgttacag tccggtgcgt ttttggtttt ttgaaagtgc
     5101 gtcttcagag cgcttttggt tttcaaaagc gctctgaagt tcctatactt tctagagaat
     5161 aggaacttcg gaataggaac ttcaaagcgt ttccgaaaac gagcgcttcc gaaaatgcaa
     5221 cgcgagctgc gcacatacag ctcactgttc acgtcgcacc tatatctgcg tgttgcctgt
     5281 atatatatat acatgagaag aacggcatag tgcgtgttta tgcttaaatg cgtacttata
     5341 tgcgtctatt tatgtaggat gaaaggtagt ctagtacctc ctgtgatatt atcccattcc
     5401 atgcggggta tcgtatgctt ccttcagcac taccctttag ctgttctata tgctgccact
     5461 cctcaattgg attagtctca tccttcaatg ctatcatttc ctttgatatt ggatcatcta
     5521 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
     5581 tc
//
