LOCUS       sequence-369732-        5623 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Markerless yeast integration Vector at Int.2 locus to used in the 
            Dueber YTK and Ellis MYT systems.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5623)
  AUTHORS   Shaw WM, Khalil AS, Ellis T
  TITLE     A Multiplex MoClo Toolkit for Extensive and Flexible Engineering of 
            Saccharomyces cerevisiae.
  JOURNAL   ACS Synth Biol. 2023 Nov 6. doi: 10.1021/acssynbio.3c00423.
  PUBMED    37930278
REFERENCE   2  (bases 1 to 5623)
  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"; journalName: "ACS Synth 
            Biol. 2023 Nov 6. doi: 10.1021/acssynbio.3c00423."
COMMENT     Sequence Label: pMYT076
FEATURES             Location/Qualifiers
     source          1..5623
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(1627..1654)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(1670..1741)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     RBS             2455..2466
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     rep_origin      complement(3447..4035)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3527..3546)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4249..4268)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4260..5075
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             4260..5075
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4333..4352)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 gcacgactgc cgttgtcgaa gccaccaagt ccaaagtaac ggcatggact catgtcttcg
       61 actgaagaca ttgtaagctc tgcagtgaca ctagtgtact gcttgagacg tagccagctg
      121 agacccagac gagcaccaaa tacccgctgg actgttgtga tatgccatat atgcagtggt
      181 gttcgccggc gctggttccg gctgtcttgc ttagttgtga gtcgccaggc agtcatcggt
      241 atgatctgta catgattcgt cagtgtgagc accactgacg agcagatttc gcggccgcag
      301 tacagctcta ggtctataaa aaaaaaaatt ggttaaagaa aaaatataca ggtttgtata
      361 tagaatcatt aattaaaaag ataatagtat aaaacacatc agataaacaa aagtttgccc
      421 aataaaatta tattttccat caaccagcta cttgtacaac tcatccatac caccagtaga
      481 atgtctacct tctgatcttt cgtactgttc aacaacggtg taatcttcgt tgtgagaggt
      541 gatatccaat tttctatcaa cgttataagc accaggcatt tgaactggct ttttggcctt
      601 gtaagtagtc ttaaaatcag ccaagtatct accaccgtct ttcaatctca aggccatctt
      661 aatatcacct ttcaaaacac catcctctgg atataatctt tcagtagaag cttcccatcc
      721 catagtcttt ttttgcataa caggaccatc tggtgggaaa ttagtacctc ttaacttaac
      781 cttgtagatc aaggtgccat cttccaatga agtatcttga gtaacagtaa cggcaccacc
      841 atcttcaaaa ttcatgactc tttcccactt gaaaccctct ggaaatgatt gcttataata
      901 atctgggata tcagctgggt gttttgtaaa agctcttgaa ccgtacataa attgtggaga
      961 caaaatatcc caagagaatg gcaaaggacc accttttgta accttcaatt tagcagtttg
     1021 tgtaccttcg tatggtctac cctcaccttc accctcaatt tcaaattcgt gaccgttcat
     1081 agaaccttcc atgtgaacct tgaatctcat aaattctttg atgacggctt cacctttaga
     1141 caccatttta attgtttgtg tgtgtgtgtt ttcgaaacta agttcttggg gttttaaaat
     1201 caataagaat taaaatagaa taactataaa ataagaagtg taaaggagtt atagttttat
     1261 gagtttatga tcgatagttg tcgtctttat atatttatta atcaaacagg gggatgattt
     1321 gtggaaactg gagacaacct ttttttcagt tttttccaac ccagagagtt agaaggtaat
     1381 agaaggtgtg agaaaataga cagacacgta ggtcgattgc cttgtgtcag ctttttgctc
     1441 cagtcagttt gcatcactcc attgattttt ttcaggaagc cagattaaaa agcgcgatcc
     1501 gagatattta gtggatgctg gaaaagggac cgctcaccca gacattcgga atgtggcctg
     1561 tggctgccgt aacccgctca atgagaagtc gggttactaa tggcaggtga aagatgaatg
     1621 tataaacgca gaaaggccca cccgaaggtg agccagtgtg actctagtag agagcgttca
     1681 ccgacaaaca acagataaaa cgaaaggccc agtctttcga ctgagccttt cgttttattt
     1741 gatgcctggt cacttataca gttcgtccat gccgccggtg ctgtggcgac cctcgctacg
     1801 ctcatattgc tcaaccacgg tataatcctc gttgtggctg gtaatatcca gtttacggtc
     1861 aacgttgtac gcacccggca tttgcaccgg tttcttcgct ttataggtgg tcttaaaatc
     1921 cgccaggtaa cggccaccgt ccttcagacg cagcgccatt ttgatatcac ccttcagcac
     1981 gccatcttcc ggatacagac gctcggtgct cgcttcccaa cccatcgttt tcttctgcat
     2041 aaccggacca tccggcggaa agttggtgcc acgcagtttc accttgtaaa tcagggtgcc
     2101 atcctccagg ctggtgtcct gggtaacggt caccgcgcca ccatcttcga agttcataac
     2161 acgctcccat ttaaaacctt ccgggaagct ttgcttatag taatccggaa tgtccgccgg
     2221 gtgtttggta aacgcacggc tgccatacat gaactgcggg ctcaggatgt cccagctgaa
     2281 cggcagcggg ccacccttgg tcactttcag cttcgcggtc tgggtgcctt cgtacggacg
     2341 accctcgcct tcaccctcga tctcgaactc gtgaccgttc atgctaccct ccatgtgaac
     2401 cttaaaacgc ataaactcct tgataaccgc ctcgcctttg ctaaccatct agtatttctc
     2461 ctctttctct agtagctaag atccgaacct gccgtttctt gagttgccgc gatgttaaga
     2521 aaacattcat aaattaaatg tgaattgccg cacacattat taaataagat ttacaaaatg
     2581 ttcaaaatga cgcatgaaat cacgtttcac tttcgcggcc gcgtattgcg acgaattgcc
     2641 acgttgtcgc agtgtctgta ctgtgggtta atgtctgtga tcgccgtcgt gagttcgtgt
     2701 catccgcgca tgattcagat ggtcctggag atcgttgctt gtaccggctg tttgtggctc
     2761 taccagccaa cgcacttttg accacagggt ctcacgttat gccgtctcat cagcttctct
     2821 agaagtcgaa ttcgggatag gggagacaag aacctgtcac ttctcgttta atatctttaa
     2881 tcgcatcatc tatagttcct aattctgcat tcctaacact gtagcctatc cagcaaaatc
     2941 attcccattt tctctttata tgtgcctaga ttttaaaatt ttgcttaaga tgatttacat
     3001 tgagggggga aacttgcttt atccgctcct ccaattgatt caagaagtat gggcgctgac
     3061 ctaccatctc aatttcttgt ttcactcggg aattgtcact gagatagtaa atgtctggag
     3121 aatcctttgc aacaggagat ttcaaattta tattctttgt gttcgcattc gcttcccgtc
     3181 tgttactcgt tcccttacca ttctattatg cattgcaggt ctgccaagta aaaaaaaaaa
     3241 taccttggtg tacacaccac ccatacatta tctgacaagg aaacaaaagt aatgacagta
     3301 aattgaagaa cttgtgatga gccaaaaaga agaatgtgtg cggccgcacg gttatccaca
     3361 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     3421 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     3481 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     3541 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     3601 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     3661 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     3721 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     3781 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     3841 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     3901 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     3961 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     4021 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     4081 gaaaactcac gttaagggat tttggtcatg actgatcctt caactcagca aaagttcgat
     4141 ttattcaaca aagccacgtt gtgtctcaaa atctctgatg ttacattgca caagataaaa
     4201 atatatcatc atgaacaata aaactgtctg cttacataaa cagtaataca aggggtgtta
     4261 tgagccatat tcaacgggaa acgtcttgct ccaggccgcg attaaattcc aacatggatg
     4321 ctgatttata tgggtataaa tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct
     4381 atcgattgta tgggaagccc gatgcgccag agttgtttct gaaacatggc aaaggtagcg
     4441 ttgccaatga tgttacagat gagatggtca gactaaactg gctgacggaa tttatgcctc
     4501 ttccgaccat caagcatttt atccgtactc ctgatgatgc atggttactc accactgcga
     4561 tccccgggaa aacagcattc caggtattag aagaatatcc tgattcaggt gaaaatattg
     4621 ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt
     4681 ttaacagcga tcgcgtattt cgcctcgctc aggcgcaatc acgaatgaat aacggtttgg
     4741 ttgatgcgag tgattttgat gacgagcgta atggctggcc tgttgaacaa gtctggaaag
     4801 aaatgcataa gcttttgcca ttctcaccgg attcagtcgt cactcatggt gatttctcac
     4861 ttgataacct tatttttgac gaggggaaat taataggttg tattgatgtt ggacgagtcg
     4921 gaatcgcaga ccgataccag gatcttgcca tcctatggaa ctgcctcggt gagttttctc
     4981 cttcattaca gaaacggctt tttcaaaaat atggtattga taatcctgat atgaataaat
     5041 tgcagtttca tttgatgctc gatgagtttt tctaatcaga attggttaat tggttgtaac
     5101 acgcggccgc tttcgtcaca tgttcaacta aattccaaac agttgaactt tttgttctac
     5161 tgatcttgta attcatgctt ccttctatgc aacacgccat actaatttag tagaatgaac
     5221 agtaactggt gaacgatgac tctgtagtag aatactataa aaaggtggca tcgtataaag
     5281 gtgccttctt tgacccttgg caggtagggc ggcaacggca caatgatatt ttatttaaca
     5341 aaaagatacg ctaacaccgc atctgaacac gtaatactac tatatgactt ggaacaaatg
     5401 gctaatcaat aatcacatat gttcgaccgc cgcaggaaaa gagaaaagaa agcagtttct
     5461 acaagcaaga acttggaaat acagacaaac ctccagtact ttattaggat tgtccattcc
     5521 tcacccagat atgttaaatt cgataccaat gtgttttgat tcacgcgcag aaagaaggac
     5581 tttccctgct ctgctcggca aaaacataca cgttgtcgac cga
//
