LOCUS       sequence-253015-        3504 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 1 destination vector for a transcription unit, pathway 
            position 1. 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-5a
FEATURES             Location/Qualifiers
     source          1..3504
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     51..70
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     protein_bind    277..310
                     /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             711..1016
                     /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     895..914
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(1088..1121)
                     /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(1591..1608)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1591..1607)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1706..1723)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1877..2465)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1957..1976)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2587..3396)
                     /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(3397..3488)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaccaattaa ccaattctga acattatcgc gagcccattt atacctgaat atggctcata
       61 acaccccttg tttgcctggc ggcagtagcg cggtggtccc acctgacccc atgccgaact
      121 cagaagtgaa acgccgtagc gccgatggta gtgtggggac tccccatgcg agagtaggga
      181 actgccaggc atcaaataaa acgaaaggct cagtcgaaag actgggcctt tcgcccgggc
      241 taattagggg gtgtcgccct tattcgactc tatagtgaag ttcctattct ctagaaagta
      301 taggaacttc tgaagtgggg aagcgacttc caatcgcttt gcatatccag taccacaccc
      361 acaggcgttt gtgcggagac cggcttacta aaagccagat aacagtatgc atatttgcgc
      421 gctgattttt gcggtataag aatatatact gatatgtata cccgaagtat gtcaaaaaga
      481 ggtatgctat gaagcagcgt attacagtga cagttgacag cgacagctat cagttgctca
      541 aggcatatat gatgtcaata tctccggtct ggtaagcaca accatgcaga atgaagcccg
      601 tcgtctgcgt gccgaacgct ggaaagcgga aaatcaggaa gggatggctg aggtcgcccg
      661 gtttattgaa atgaacggct cttttgctga cgagaacagg ggctggtgaa atgcagttta
      721 aggtttacac ctataaaaga gagagccgtt atcgtctgtt tgtggatgta cagagtgata
      781 ttattgacac gcccgggcga cggatggtga tccccctggc cagtgcacgt ctgctgtcag
      841 ataaagtctc ccgtgaactt tacccggtgg tgcatatcgg ggatgaaagc tggcgcatga
      901 tgaccaccga tatggccagt gtgccggttt ccgttatcgg ggaagaagtg gctgatctca
      961 gccaccgcga aaatgacatc aaaaacgcca ttaacctgat gttctgggga atataaggtc
     1021 tcgcctcttg cccatcgaac gtacaagtac tcctctgttc tctccttcct ttgctttaaa
     1081 atgaagtgaa gttcctatac tttctagaga ataggaactt ctatagtgag tcgaataagg
     1141 gcgacacaaa atttattcta aatgcataat aaatactgat aacatcttat agtttgtatt
     1201 atattttgta ttatcgttga catgtataat tttgatatca aaaactgatt ttccctttat
     1261 tattttcgag atttattttc ttaattctct ttaacaaact agaaatattg tatatacaaa
     1321 aaatcataaa taatagatga atagtttaat tataggtgtt catcaatcga aaaagcaacg
     1381 tatcttattt aaagtgcgtt gcttttttct catttataag gttaaataat tctcatatat
     1441 caagcaaagt gacaggcgcc cttaaatatt ctgacaaatg ctctttccct aaactccccc
     1501 cataaaaaaa cccgccgaag cgggttttta cgttatttgc ggattaacga ttactcgtta
     1561 tcagaaccgc ccagggggcc cgagcttaag actggccgtc gttttacaac acagaaagag
     1621 tttgtagaaa cgcaaaaagg ccatccgtca ggggccttct gcttagtttg atgcctggca
     1681 gttccctact ctcgccttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     1741 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     1801 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     1861 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     1921 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     1981 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     2041 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     2101 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     2161 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     2221 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     2281 tcttgaagtg gtgggctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     2341 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     2401 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     2461 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gacgcgcgcg
     2521 taactcacgt taagggattt tggtcatgag cttgcgccgt cccgtcaagt cagcgtaatg
     2581 ctctgcttag aaaaactcat cgagcatcaa atgaaattgc aatttattca tatcaggatt
     2641 atcaatacca tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca
     2701 gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat
     2761 acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt
     2821 gacgactgaa tccggtgaga atggcaaaag tttatgcatt tctttccaga cttgttcaac
     2881 aggccagcca ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg
     2941 tgattgcgcc tgagcgaggc gaaatacgcg atcgctgtta aaaggacaat tacaaacagg
     3001 aatcgagtgc aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc
     3061 aggatattct tctaatacct ggaacgctgt ttttccgggg atcgcagtgg tgagtaacca
     3121 tgcatcatca ggagtacgga taaaatgctt gatggtcgga agtggcataa attccgtcag
     3181 ccagtttagt ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt
     3241 cagaaacaac tctggcgcat cgggcttccc atacaagcga tagattgtcg cacctgattg
     3301 cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa
     3361 tcgcggcctc gacgtttccc gttgaatatg gctcatattc ttcctttttc aatattattg
     3421 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     3481 taaacaaata ggggtcagtg ttac
//
