LOCUS       sequence-253022-        3504 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 1 destination vector for a transcription unit, pathway 
            position 5. 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-de
FEATURES             Location/Qualifiers
     source          1..3504
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     27..44
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     142..159
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     143..159
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    629..662
                     /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             complement(734..1039)
                     /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     complement(836..855)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(1440..1473)
                     /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(1680..1699)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     promoter        1766..1857
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1858..2667
                     /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"
     rep_origin      2789..3377
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3278..3297
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaaggcgaga
       61 gtagggaact gccaggcatc aaactaagca gaaggcccct gacggatggc ctttttgcgt
      121 ttctacaaac tctttctgtg ttgtaaaacg acggccagtc ttaagctcgg gccccctggg
      181 cggttctgat aacgagtaat cgttaatccg caaataacgt aaaaacccgc ttcggcgggt
      241 ttttttatgg ggggagttta gggaaagagc atttgtcaga atatttaagg gcgcctgtca
      301 ctttgcttga tatatgagaa ttatttaacc ttataaatga gaaaaaagca acgcacttta
      361 aataagatac gttgcttttt cgattgatga acacctataa ttaaactatt catctattat
      421 ttatgatttt ttgtatatac aatatttcta gtttgttaaa gagaattaag aaaataaatc
      481 tcgaaaataa taaagggaaa atcagttttt gatatcaaaa ttatacatgt caacgataat
      541 acaaaatata atacaaacta taagatgtta tcagtattta ttatgcattt agaataaatt
      601 ttgtgtcgcc cttattcgac tcactataga agttcctatt ctctagaaag tataggaact
      661 tcacttcatt ttaggtacaa caagcacgac cgagcatatg ggaaggatgt ttgtggttat
      721 ttgaggcgag accttatatt ccccagaaca tcaggttaat ggcgtttttg atgtcatttt
      781 cgcggtggct gagatcagcc acttcttccc cgataacgga aaccggcaca ctggccatat
      841 cggtggtcat catgcgccag ctttcatccc cgatatgcac caccgggtaa agttcacggg
      901 agactttatc tgacagcaga cgtgcactgg ccagggggat caccatccgt cgcccgggcg
      961 tgtcaataat atcactctgt acatccacaa acagacgata acggctctct cttttatagg
     1021 tgtaaacctt aaactgcatt tcaccagccc ctgttctcgt cagcaaaaga gccgttcatt
     1081 tcaataaacc gggcgacctc agccatccct tcctgatttt ccgctttcca gcgttcggca
     1141 cgcagacgac gggcttcatt ctgcatggtt gtgcttacca gaccggagat attgacatca
     1201 tatatgcctt gagcaactga tagctgtcgc tgtcaactgt cactgtaata cgctgcttca
     1261 tagcatacct ctttttgaca tacttcgggt atacatatca gtatatattc ttataccgca
     1321 aaaatcagcg cgcaaatatg catactgtta tctggctttt agtaagccgg tctccgcacg
     1381 cggaatattg gcggaacgga cacacgtgga tacaaacctg gacaacgttc cccacttcag
     1441 aagttcctat actttctaga gaataggaac ttcactatag agtcgaataa gggcgacacc
     1501 ccctaattag cccgggcgaa aggcccagtc tttcgactga gcctttcgtt ttatttgatg
     1561 cctggcagtt ccctactctc gcatggggag tccccacact accatcggcg ctacggcgtt
     1621 tcacttctga gttcggcatg gggtcaggtg ggaccaccgc gctactgccg ccaggcaaac
     1681 aaggggtgtt atgagccata ttcaggtata aatgggctcg cgataatgtt cagaattggt
     1741 taattggttg taacactgac ccctatttgt ttatttttct aaatacattc aaatatgtat
     1801 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagaatatg
     1861 agccatattc aacgggaaac gtcgaggccg cgattaaatt ccaacatgga tgctgattta
     1921 tatgggtata aatgggctcg cgataatgtc gggcaatcag gtgcgacaat ctatcgcttg
     1981 tatgggaagc ccgatgcgcc agagttgttt ctgaaacatg gcaaaggtag cgttgccaat
     2041 gatgttacag atgagatggt cagactaaac tggctgacgg aatttatgcc acttccgacc
     2101 atcaagcatt ttatccgtac tcctgatgat gcatggttac tcaccactgc gatccccgga
     2161 aaaacagcgt tccaggtatt agaagaatat cctgattcag gtgaaaatat tgttgatgcg
     2221 ctggcagtgt tcctgcgccg gttgcactcg attcctgttt gtaattgtcc ttttaacagc
     2281 gatcgcgtat ttcgcctcgc tcaggcgcaa tcacgaatga ataacggttt ggttgatgcg
     2341 agtgattttg atgacgagcg taatggctgg cctgttgaac aagtctggaa agaaatgcat
     2401 aaacttttgc cattctcacc ggattcagtc gtcactcatg gtgatttctc acttgataac
     2461 cttatttttg acgaggggaa attaataggt tgtattgatg ttggacgagt cggaatcgca
     2521 gaccgatacc aggatcttgc catcctatgg aactgcctcg gtgagttttc tccttcatta
     2581 cagaaacggc tttttcaaaa atatggtatt gataatcctg atatgaataa attgcaattt
     2641 catttgatgc tcgatgagtt tttctaagca gagcattacg ctgacttgac gggacggcgc
     2701 aagctcatga ccaaaatccc ttaacgtgag ttacgcgcgc gtcgttccac tgagcgtcag
     2761 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     2821 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     2881 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     2941 tagtgtagcc gtagttagcc caccacttca agaactctgt agcaccgcct acatacctcg
     3001 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     3061 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     3121 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     3181 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     3241 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     3301 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     3361 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     3421 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     3481 ccgcctttga gtgagctgat accg
//
