LOCUS       sequence-351133-        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"
     RBS             882..893
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      1607..1678
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1694..1721
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(3896..4711)
                     /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             complement(3896..4711)
                     /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     4619..4638
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4703..4722
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4936..5524
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5425..5444
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gcggccgcac acattcttct ttttggctca tcacaagttc ttcaatttac tgtcattact
       61 tttgtttcct tgtcagataa tgtatgggtg gtgtgtacac caaggtattt ttttttttac
      121 ttggcagacc tgcaatgcat aatagaatgg taagggaacg agtaacagac gggaagcgaa
      181 tgcgaacaca aagaatataa atttgaaatc tcctgttgca aaggattctc cagacattta
      241 ctatctcagt gacaattccc gagtgaaaca agaaattgag atggtaggtc agcgcccata
      301 cttcttgaat caattggagg agcggataaa gcaagtttcc cccctcaatg taaatcatct
      361 taagcaaaat tttaaaatct aggcacatat aaagagaaaa tgggaatgat tttgctggat
      421 aggctacagt gttaggaatg cagaattagg aactatagat gatgcgatta aagatattaa
      481 acgagaagtg acaggttctt gtctccccta tcccgaattc gacttctaga gaagctgatg
      541 agacggcata acgtgagacc ctgtggtcaa aagtgcgttg gctggtagag ccacaaacag
      601 ccggtacaag caacgatctc caggaccatc tgaatcatgc gcggatgaca cgaactcacg
      661 acggcgatca cagacattaa cccacagtac agacactgcg acaacgtggc aattcgtcgc
      721 aatacgcggc cgcgaaagtg aaacgtgatt tcatgcgtca ttttgaacat tttgtaaatc
      781 ttatttaata atgtgtgcgg caattcacat ttaatttatg aatgttttct taacatcgcg
      841 gcaactcaag aaacggcagg ttcggatctt agctactaga gaaagaggag aaatactaga
      901 tggttagcaa aggcgaggcg gttatcaagg agtttatgcg ttttaaggtt cacatggagg
      961 gtagcatgaa cggtcacgag ttcgagatcg agggtgaagg cgagggtcgt ccgtacgaag
     1021 gcacccagac cgcgaagctg aaagtgacca agggtggccc gctgccgttc agctgggaca
     1081 tcctgagccc gcagttcatg tatggcagcc gtgcgtttac caaacacccg gcggacattc
     1141 cggattacta taagcaaagc ttcccggaag gttttaaatg ggagcgtgtt atgaacttcg
     1201 aagatggtgg cgcggtgacc gttacccagg acaccagcct ggaggatggc accctgattt
     1261 acaaggtgaa actgcgtggc accaactttc cgccggatgg tccggttatg cagaagaaaa
     1321 cgatgggttg ggaagcgagc accgagcgtc tgtatccgga agatggcgtg ctgaagggtg
     1381 atatcaaaat ggcgctgcgt ctgaaggacg gtggccgtta cctggcggat tttaagacca
     1441 cctataaagc gaagaaaccg gtgcaaatgc cgggtgcgta caacgttgac cgtaaactgg
     1501 atattaccag ccacaacgag gattataccg tggttgagca atatgagcgt agcgagggtc
     1561 gccacagcac cggcggcatg gacgaactgt ataagtgacc aggcatcaaa taaaacgaaa
     1621 ggctcagtcg aaagactggg cctttcgttt tatctgttgt ttgtcggtga acgctctcta
     1681 ctagagtcac actggctcac cttcgggtgg gcctttctgc gtttatacat tcatctttca
     1741 cctgccatta gtaacccgac ttctcattga gcgggttacg gcagccacag gccacattcc
     1801 gaatgtctgg gtgagcggtc ccttttccag catccactaa atatctcgga tcgcgctttt
     1861 taatctggct tcctgaaaaa aatcaatgga gtgatgcaaa ctgactggag caaaaagctg
     1921 acacaaggca atcgacctac gtgtctgtct attttctcac accttctatt accttctaac
     1981 tctctgggtt ggaaaaaact gaaaaaaagg ttgtctccag tttccacaaa tcatccccct
     2041 gtttgattaa taaatatata aagacgacaa ctatcgatca taaactcata aaactataac
     2101 tcctttacac ttcttatttt atagttattc tattttaatt cttattgatt ttaaaacccc
     2161 aagaacttag tttcgaaaac acacacacac aaacaattaa aatggtgtct aaaggtgaag
     2221 ccgtcatcaa agaatttatg agattcaagg ttcacatgga aggttctatg aacggtcacg
     2281 aatttgaaat tgagggtgaa ggtgagggta gaccatacga aggtacacaa actgctaaat
     2341 tgaaggttac aaaaggtggt cctttgccat tctcttggga tattttgtct ccacaattta
     2401 tgtacggttc aagagctttt acaaaacacc cagctgatat cccagattat tataagcaat
     2461 catttccaga gggtttcaag tgggaaagag tcatgaattt tgaagatggt ggtgccgtta
     2521 ctgttactca agatacttca ttggaagatg gcaccttgat ctacaaggtt aagttaagag
     2581 gtactaattt cccaccagat ggtcctgtta tgcaaaaaaa gactatggga tgggaagctt
     2641 ctactgaaag attatatcca gaggatggtg ttttgaaagg tgatattaag atggccttga
     2701 gattgaaaga cggtggtaga tacttggctg attttaagac tacttacaag gccaaaaagc
     2761 cagttcaaat gcctggtgct tataacgttg atagaaaatt ggatatcacc tctcacaacg
     2821 aagattacac cgttgttgaa cagtacgaaa gatcagaagg tagacattct actggtggta
     2881 tggatgagtt gtacaagtag ctggttgatg gaaaatataa ttttattggg caaacttttg
     2941 tttatctgat gtgttttata ctattatctt tttaattaat gattctatat acaaacctgt
     3001 atattttttc tttaaccaat tttttttttt atagacctag agctgtactg cggccgcgaa
     3061 atctgctcgt cagtggtgct cacactgacg aatcatgtac agatcatacc gatgactgcc
     3121 tggcgactca caactaagca agacagccgg aaccagcgcc ggcgaacacc actgcatata
     3181 tggcatatca caacagtcca gcgggtattt ggtgctcgtc tgggtctcag ctggctacgt
     3241 ctcaagcagt acactagtgt cactgcagag cttacaatgt cttcagtcga agacatgagt
     3301 ccatgccgtt actttggact tggtggcttc gacaacggca gtcgtgctcg gtcgacaacg
     3361 tgtatgtttt tgccgagcag agcagggaaa gtccttcttt ctgcgcgtga atcaaaacac
     3421 attggtatcg aatttaacat atctgggtga ggaatggaca atcctaataa agtactggag
     3481 gtttgtctgt atttccaagt tcttgcttgt agaaactgct ttcttttctc ttttcctgcg
     3541 gcggtcgaac atatgtgatt attgattagc catttgttcc aagtcatata gtagtattac
     3601 gtgttcagat gcggtgttag cgtatctttt tgttaaataa aatatcattg tgccgttgcc
     3661 gccctacctg ccaagggtca aagaaggcac ctttatacga tgccaccttt ttatagtatt
     3721 ctactacaga gtcatcgttc accagttact gttcattcta ctaaattagt atggcgtgtt
     3781 gcatagaagg aagcatgaat tacaagatca gtagaacaaa aagttcaact gtttggaatt
     3841 tagttgaaca tgtgacgaaa gcggccgcgt gttacaacca attaaccaat tctgattaga
     3901 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
     3961 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
     4021 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
     4081 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
     4141 ccggtgagaa tggcaaaagc ttatgcattt ctttccagac ttgttcaaca ggccagccat
     4201 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
     4261 gagcgaggcg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
     4321 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
     4381 ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcatcatcag
     4441 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
     4501 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
     4561 ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat
     4621 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctgg
     4681 agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag
     4741 cagacagttt tattgttcat gatgatatat ttttatcttg tgcaatgtaa catcagagat
     4801 tttgagacac aacgtggctt tgttgaataa atcgaacttt tgctgagttg aaggatcagt
     4861 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     4921 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     4981 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     5041 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     5101 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     5161 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     5221 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     5281 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     5341 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     5401 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     5461 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     5521 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     5581 catgttcttt cctgcgttat cccctgattc tgtggataac cgt
//
