LOCUS       sequence-253018-        3504 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 1 destination vector for a transcription unit, pathway 
            position 3. Clone a transcription unit in with BsaI..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3504)
  AUTHORS   Young EM, Zhao Z, Gielesen BEM, Wu L, Benjamin Gordon D, Roubos JA, 
            Voigt CA
  TITLE     Iterative algorithm-guided design of massive strain libraries, 
            applied to itaconic acid production in yeast.
  JOURNAL   Metab Eng. 2018 Jul;48:33-43. doi: 10.1016/j.ymben.2018.05.002. Epub
            2018 May 9.
  PUBMED    29753070
REFERENCE   2  (bases 1 to 3504)
  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: "Metab 
            Eng."; date: "2018-07"; volume: "48:33-43. doi"; pages: " 
            10.1016/j.ymben.2018.05.002. Epub 2018 May 9"
COMMENT     Sequence Label: pDSM-bc
FEATURES             Location/Qualifiers
     source          1..3504
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    194..227
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             628..933
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     812..831
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(1005..1038)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(1508..1525)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1508..1524)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1623..1640)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1794..2382)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1874..1893)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2504..3313)
                     /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="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     promoter        complement(3314..3405)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3472..3491
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 agtagcgcgg tggtcccacc tgaccccatg ccgaactcag aagtgaaacg ccgtagcgcc
       61 gatggtagtg tggggactcc ccatgcgaga gtagggaact gccaggcatc aaataaaacg
      121 aaaggctcag tcgaaagact gggcctttcg cccgggctaa ttagggggtg tcgcccttat
      181 tcgactctat agtgaagttc ctattctcta gaaagtatag gaacttctga agtggggcgg
      241 atcgatgtac acaaccgact gcacccaaac gaacacaaat cttagcagtg cggagaccgg
      301 cttactaaaa gccagataac agtatgcata tttgcgcgct gatttttgcg gtataagaat
      361 atatactgat atgtataccc gaagtatgtc aaaaagaggt atgctatgaa gcagcgtatt
      421 acagtgacag ttgacagcga cagctatcag ttgctcaagg catatatgat gtcaatatct
      481 ccggtctggt aagcacaacc atgcagaatg aagcccgtcg tctgcgtgcc gaacgctgga
      541 aagcggaaaa tcaggaaggg atggctgagg tcgcccggtt tattgaaatg aacggctctt
      601 ttgctgacga gaacaggggc tggtgaaatg cagtttaagg tttacaccta taaaagagag
      661 agccgttatc gtctgtttgt ggatgtacag agtgatatta ttgacacgcc cgggcgacgg
      721 atggtgatcc ccctggccag tgcacgtctg ctgtcagata aagtctcccg tgaactttac
      781 ccggtggtgc atatcgggga tgaaagctgg cgcatgatga ccaccgatat ggccagtgtg
      841 ccggtttccg ttatcgggga agaagtggct gatctcagcc accgcgaaaa tgacatcaaa
      901 aacgccatta acctgatgtt ctggggaata taaggtctcg cctcacgctt tccggcatct
      961 tccagaccac agtatatcca tccgcctcct gttgaaaatg aagtgaagtt cctatacttt
     1021 ctagagaata ggaacttcta tagtgagtcg aataagggcg acacaaaatt tattctaaat
     1081 gcataataaa tactgataac atcttatagt ttgtattata ttttgtatta tcgttgacat
     1141 gtataatttt gatatcaaaa actgattttc cctttattat tttcgagatt tattttctta
     1201 attctcttta acaaactaga aatattgtat atacaaaaaa tcataaataa tagatgaata
     1261 gtttaattat aggtgttcat caatcgaaaa agcaacgtat cttatttaaa gtgcgttgct
     1321 tttttctcat ttataaggtt aaataattct catatatcaa gcaaagtgac aggcgccctt
     1381 aaatattctg acaaatgctc tttccctaaa ctccccccat aaaaaaaccc gccgaagcgg
     1441 gtttttacgt tatttgcgga ttaacgatta ctcgttatca gaaccgccca gggggcccga
     1501 gcttaagact ggccgtcgtt ttacaacaca gaaagagttt gtagaaacgc aaaaaggcca
     1561 tccgtcaggg gccttctgct tagtttgatg cctggcagtt ccctactctc gccttccgct
     1621 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     1681 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     1741 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     1801 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     1861 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     1921 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     1981 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     2041 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     2101 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     2161 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg ggctaactac
     2221 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     2281 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     2341 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     2401 tctacggggt ctgacgctca gtggaacgac gcgcgcgtaa ctcacgttaa gggattttgg
     2461 tcatgagctt gcgccgtccc gtcaagtcag cgtaatgctc tgcttagaaa aactcatcga
     2521 gcatcaaatg aaattgcaat ttattcatat caggattatc aataccatat ttttgaaaaa
     2581 gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct
     2641 ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt
     2701 caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg
     2761 gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat
     2821 caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcgaggcgaa
     2881 atacgcgatc gctgttaaaa ggacaattac aaacaggaat cgagtgcaac cggcgcagga
     2941 acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct aatacctgga
     3001 acgctgtttt tccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa
     3061 aatgcttgat ggtcggaagt ggcataaatt ccgtcagcca gtttagtctg accatctcat
     3121 ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg
     3181 gcttcccata caagcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt
     3241 tatacccata taaatcagca tccatgttgg aatttaatcg cggcctcgac gtttcccgtt
     3301 gaatatggct catattcttc ctttttcaat attattgaag catttatcag ggttattgtc
     3361 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gtcagtgtta
     3421 caaccaatta accaattctg aacattatcg cgagcccatt tatacctgaa tatggctcat
     3481 aacacccctt gtttgcctgg cggc
//
